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Frameshift mutation

A frameshift mutation is a type of genetic mutation resulting from the insertion or deletion of a number of nucleotides in a DNA sequence that is not divisible by three, thereby altering the reading frame of the genetic code and leading to a completely different translation of the genetic information from the mutation site onward. This disruption was first experimentally demonstrated in 1961 by Francis Crick, Sydney Brenner, and colleagues through studies on bacteriophage T4, where combinations of insertions and deletions restored function, confirming the triplet nature of the genetic code. Mechanistically, the mRNA is read in codons of three nucleotides during translation; an insertion or deletion of one or two bases shifts this frame, changing all subsequent codons and typically producing a garbled amino acid sequence in the resulting protein, often culminating in a premature stop codon that truncates the polypeptide. Such mutations are usually deleterious, rendering proteins nonfunctional and contributing to a wide array of genetic disorders, including Crohn's disease via a frameshift in the NOD2 gene, certain forms of cystic fibrosis, and Tay-Sachs disease. Despite their predominantly harmful effects, frameshift mutations play a role in evolutionary processes by potentially generating novel protein functions or regulatory elements, though they are far more often associated with loss-of-function phenotypes in human health.

Background and Fundamentals

Definition and characteristics

A frameshift mutation is a type of genetic mutation caused by the insertion or deletion of a number of in that is not divisible by three, thereby shifting the of the and altering the translation of the subsequent codons into . This shift occurs because the is read in triplets, known as codons, each specifying a particular or stop signal during protein synthesis. The concept of frameshift mutations was first experimentally demonstrated in 1961 by , , and colleagues through their work on proflavin-induced mutations in T4, which provided key evidence for the triplet nature of the and the disruptive effects of such shifts. Key characteristics of frameshift mutations include their tendency to produce a cascade of incorrect substitutions downstream from the mutation site, often introducing a premature that truncates the protein. These alterations typically result in non-functional or loss-of-function proteins, as the changed sequence disrupts the protein's structure and activity. Frameshift mutations differ from in-frame insertions or deletions, which involve multiples of three nucleotides and thus add or remove whole codons without altering the downstream reading frame, potentially preserving partial protein function depending on the affected residues.

The genetic code and reading frames

The outlines the unidirectional flow of genetic information within cells, proceeding from DNA to (mRNA) through transcription, and subsequently from mRNA to proteins via translation. This framework, first proposed by , establishes that genetic instructions encoded in DNA are copied into mRNA, which serves as the template for synthesizing polypeptide chains that fold into functional proteins. The genetic code operates on a triplet basis, where sequences of three consecutive nucleotides, known as codons, specify individual amino acids or termination signals during translation. With four possible nucleotides (adenine, cytosine, guanine, and uracil in mRNA), there are 64 possible codons (4³ = 64), which encode the 20 standard amino acids and three stop signals (UAA, UAG, and UGA) that halt protein synthesis. This degeneracy, or redundancy, means most amino acids are represented by multiple codons, allowing for flexibility while minimizing the impact of certain mutations. The code is read in a non-overlapping manner by ribosomes, which scan mRNA from a defined starting point to assemble amino acids in the precise order dictated by successive codons. In any given DNA or mRNA sequence, three possible reading frames exist, corresponding to the three alternative ways to group nucleotides into triplets starting from different positions (e.g., positions 1-3, 2-4, or 3-5). The correct reading frame, often termed the open reading frame (ORF), begins at an initiation codon (typically AUG in mRNA, coding for methionine) and extends continuously to a stop codon without intervening stops, ensuring accurate translation of the intended protein sequence. Maintenance of this frame is essential, as any disruption—such as the insertion or deletion of nucleotides not in multiples of three—shifts the reading frame, misaligning all subsequent codons and typically resulting in a completely altered amino acid sequence downstream, often culminating in a premature stop codon. Frameshift mutations exemplify this vulnerability, scrambling the genetic message beyond the alteration site.

Causes

Genetic mechanisms

Frameshift mutations primarily arise from small insertions or deletions (indels) of that are not multiples of three, disrupting the of the . These indels often occur during through a process known as replication slippage, where temporarily dissociates from the template strand and reanneals out of register, particularly in repetitive DNA sequences such as microsatellites. Microsatellites, consisting of short tandem repeats, are especially prone to this slippage because the repetitive nature facilitates misalignment, leading to the addition or loss of repeat units that manifest as frameshifts. DNA polymerase errors contribute significantly to frameshift generation, as replicative polymerases like Pol δ and Pol ε exhibit varying during , with lower accuracy in repetitive regions where may fail to correct slippage-induced mismatches. Error-prone polymerases, such as those involved in translesion , further exacerbate rates by incorporating imprecisely during replication stress. Deficiencies in mismatch repair (MMR) pathways amplify these errors, as MMR normally excises and replaces mismatched bases or small loops formed by slippage; impaired MMR, as seen in conditions like Lynch syndrome caused by mutations in MLH1 or MSH2 genes, results in a markedly elevated frequency of frameshift , particularly in loci. Recombination processes also generate frameshifts through error-prone mechanisms. Unequal crossing-over during in regions with tandem repeats can produce net insertions or deletions by misaligning homologous chromosomes, shifting the reading frame in the recombinant products. Similarly, (NHEJ), a pathway for repairing double-strand breaks, often introduces small indels at junctions due to imprecise of blunt or overhanging ends, converting breaks into frameshift mutations when occurring within coding sequences. Frameshift mutations occur at higher frequencies in non-coding regions of the compared to coding exons, where strong purifying selection limits their fixation due to the predominantly deleterious effects on protein function. While most frameshifts are harmful, reducing by producing truncated or aberrant proteins, they can play an evolutionary role by generating , as evidenced by compensatory frameshifts that restore reading frames and contribute to protein in vertebrates and . Environmental factors can amplify these intrinsic mechanisms by inducing replication , but the core processes remain rooted in cellular replication and repair fidelity.

Environmental and mutagenic factors

Chemical mutagens, particularly intercalating agents such as acridines (e.g., proflavin), induce frameshift mutations by inserting between DNA base pairs, which distorts the DNA helix and promotes the addition or deletion of nucleotides during replication. These agents stabilize slipped DNA structures, leading to misalignment and indel formation in repetitive sequences. Physical mutagens like (UV) radiation generate cyclobutane , especially dimers, which block replication forks and trigger error-prone translesion synthesis, resulting in insertions or deletions that cause frameshifts. produces double-strand breaks that, during repair, often lead to small insertions or deletions, manifesting as frameshift mutations in surviving cells. These physical agents are prevalent in environmental exposures such as and cosmic rays. Biological factors, including viral integrations, can insert genetic material into host genomes, disrupting coding sequences and introducing frameshifts if the insertion length is not a multiple of three. Transposon activity similarly causes frameshift mutations through the excision or insertion of transposable elements within exons, altering the and protein function. Occupational exposure to chemical mutagens, such as certain drugs (e.g., alkylating agents), heightens risk by promoting DNA adducts that lead to formation during replication. In human exposure contexts, smoking introduces polycyclic aromatic hydrocarbons and other chemicals that elevate somatic mutation rates, including frameshifts, in lung epithelial cells through oxidative damage and replication errors. Air pollution, particularly particulate matter like PM2.5, correlates with increased somatic mutations in lung tissues by inducing chronic inflammation and DNA damage, amplifying mutation burdens in non-smokers. These environmental factors interact with genetic repair deficiencies, worsening frameshift accumulation in vulnerable populations.

Molecular Consequences

Effects on mRNA and protein synthesis

Frameshift mutations, which involve the insertion or deletion of not divisible by three in the DNA coding sequence, are faithfully transcribed by into (mRNA), resulting in a corresponding in the mRNA transcript that shifts the downstream of the site. This alteration does not directly impair the transcription process itself, such as the initiation, elongation, or termination by , but instead modifies the template sequence, leading to an mRNA with a disrupted codon for subsequent . During , the begins decoding the mRNA from the in the original until it reaches the site, after which the shifted frame causes all subsequent codons to be read incorrectly, often producing a polypeptide with an entirely different sequence. This frameshift frequently introduces a premature (such as UGA, UAA, or UAG in the new frame) shortly downstream, truncating the protein and halting synthesis prematurely. For instance, a +1 frameshift can realign the sequence to create a stop codon, preventing the full-length protein from being assembled. Many frameshift-induced premature stop codons trigger (NMD), a surveillance mechanism that identifies and degrades mRNAs with termination codons located more than 50-55 upstream of an exon-exon , thereby preventing the production of potentially harmful truncated proteins. NMD involves factors like UPF1, which recruit exonucleases to degrade the aberrant mRNA, often significantly reducing mutant transcript levels compared to wild-type, typically by more than 90% in efficient cases. This degradation significantly diminishes overall protein output from the affected . The combined effects of frameshifting and NMD typically result in a near-complete loss of functional protein from the mutated , leading to approximately 50% total protein levels from the wild-type in heterozygous cells. In genes where one functional copy is insufficient for normal dosage, this can lead to pronounced phenotypic effects due to , as the wild-type alone cannot compensate fully. Experimental studies, such as those using minigene constructs, confirm that inhibiting NMD partially restores protein expression but often yields nonfunctional products due to the underlying frameshift.

Types of resulting proteins

Frameshift mutations disrupt the of the , typically resulting in proteins that are dysfunctional due to altered sequences downstream of the site. The predominant type of aberrant protein produced is a truncated polypeptide, arising from the frequent introduction of a premature termination codon in the new reading frame. These shortened proteins often lack critical functional domains necessary for proper structure, stability, or interactions, leading to rapid degradation via pathways or inherent instability that precludes effective cellular roles. In contrast, elongated proteins emerge in rare instances where the frameshift mutation shifts the in a way that bypasses the original , permitting to extend into the 3' and generate an extended chain with a novel C-terminal sequence. Such proteins may incorporate additional that alter subcellular localization, dimerization capabilities, or regulatory motifs, though they generally exhibit diminished or absent compared to the wild-type form. This outcome is less common because most frameshifts align with in-frame stop codons earlier in the sequence. Beyond length alterations, the shifted codon usage often produces proteins prone to misfolding, where the incorrect string disrupts secondary and tertiary structures, promoting aggregation into insoluble complexes or triggering endoplasmic reticulum stress responses. These misfolded or aggregated forms can confer toxic gain-of-function properties, such as sequestering chaperone proteins or wild-type counterparts, thereby exacerbating cellular dysfunction independent of simple loss of function. The pathological impact of these mutant proteins varies by mechanism: occurs when the reduced protein levels from the mutant (approximately 50% total in heterozygotes) fall below a threshold required for normal , as the wild-type alone cannot compensate. Alternatively, dominant-negative effects arise if the aberrant protein actively with wild-type , for example, by co-assembling into heteromeric complexes that impair activity or by forming aggregates that deplete available functional monomers. Frameshift-derived truncated proteins are particularly associated with dominant-negative interference in multimeric assemblies.

Detection and Diagnosis

Molecular detection techniques

Frameshift mutations, caused by insertions or deletions of not divisible by three, are detected at the molecular level using techniques that identify alterations in or RNA. These methods focus on laboratory-based approaches to pinpoint indels, often combining direct sequencing with electrophoretic or fluorescent assays for confirmation. High-throughput strategies have become essential for genome-wide screening, while targeted tools provide precision for specific loci. DNA sequencing remains the cornerstone for detecting frameshift mutations. is widely used for targeted validation of , offering high accuracy in resolving small insertions or deletions through direct chromatogram analysis, where frameshifts appear as shifts in the sequence trace. This method is particularly effective for confirming suspected mutations in diploid genomes, with simple post-sequencing tools like wildcard searches enhancing indel identification efficiency. For broader detection, next-generation sequencing (NGS), including whole-exome sequencing (WES), enables high-throughput analysis of coding regions. WES has identified frameshift mutations in numerous studies of genetic disorders, highlighting NGS's role in scaling detection across thousands of genes. Large-scale WES efforts, such as those in , routinely catalog millions of , many of which are frameshifting, providing a comprehensive view of variant landscapes. Long-read sequencing technologies, such as (PacBio) HiFi and Oxford Nanopore, have emerged as complementary methods for accurate detection of frameshift mutations, particularly for resolving complex or repetitive indels that challenge short-read NGS. As of 2025, these approaches improve diagnostic yield by 10-15% in undiagnosed cases by providing phased variant calls and structural context, enabling better characterization of frameshift consequences in clinical . PCR-based methods offer sensitive, cost-effective screening for frameshift mutations by exploiting physicochemical changes induced by indels. Denaturing gradient (DGGE) separates PCR-amplified fragments based on melting behavior differences; insertions or deletions alter DNA stability, causing mobility shifts under denaturing conditions, with detection rates approaching 100% for small indels like those in polyadenine tracts. DGGE is especially useful for heterogeneous samples, as it distinguishes mutant from wild-type alleles even in low-abundance scenarios. Similarly, single-strand conformation polymorphism (SSCP) detects frameshifts through altered electrophoretic mobility of single-stranded DNA, where indels disrupt secondary structure folding. SSCP has successfully identified frameshift mutations in coding repeats, such as in the BAX gene, with sensitivity up to 90% under optimized conditions, making it a staple for initial mutation scanning before sequencing. These techniques are often combined with to amplify suspect regions, providing rapid preliminary evidence of frameshifts. Fluorescence-based techniques allow visualization of frameshift mutations, particularly in cellular or contexts. Fluorescent (FISH) can assess chromosomal copy number in genome editing workflows like , aiding interpretation of indel-induced disruptions, as demonstrated in cell lines where frameshift efficiency was quantified post-transfection. More commonly, reporter assays using (GFP) provide a functional readout; in these systems, an upstream or out-of-frame sequence is placed before the GFP , such that indels restore the frame and enable GFP expression, resulting in detectable . This approach has been refined for high-sensitivity detection in model organisms like , where injected mutant reporters confirm frameshift alleles in embryos with low background noise. Such assays are quantitative, allowing flow cytometry-based sorting of frameshift-positive cells. Bioinformatics tools complement wet-lab methods by predicting and analyzing frameshift mutations from sequence data. Algorithms employing , such as those in SIFT Indel, score for their likelihood to cause frameshifts and predict functional impacts by assessing changes in protein sequence downstream of the variant. (ORF) analysis tools like ORFfinder scan DNA sequences to identify disrupted ORFs, flagging potential frameshifts where would prematurely terminate or produce aberrant proteins. These computational approaches integrate with NGS pipelines, using software to detect indels via gap penalties and then applying ORF prediction to evaluate frameshift consequences, as seen in tools like TransPPMP for pathogenicity assessment. In large datasets, such methods facilitate automated filtering of frameshifting variants from millions of candidates.

Clinical diagnostic methods

Clinical diagnostic methods for frameshift mutations primarily involve targeted genetic testing in at-risk populations and routine screening programs to identify carriers or affected individuals early in life. panels utilize next-generation sequencing (NGS) to detect insertions or deletions (indels) in disease-associated genes, such as the CFTR gene for , where frameshift mutations account for about 16% of known pathogenic variants. These panels are recommended for confirmatory diagnosis following initial biochemical screens and for carrier screening, particularly in high-prevalence groups like , where testing for gene frameshifts, such as the 1278insTATC insertion responsible for Tay-Sachs disease, detects over 90% of carriers. The American College of Medical Genetics (ACMG) endorses panels covering at least 23 common CFTR variants, including indels, for broad clinical utility in diagnosing autosomal recessive disorders. Prenatal and newborn screening programs further enhance early detection of frameshift mutations through invasive and non-invasive approaches. , such as performed between 15 and 20 weeks of gestation, involves NGS analysis of fetal DNA to identify indels in genes like CFTR when both parents are known carriers, allowing for informed reproductive decisions. , standard in many countries including all U.S. states for , begins with a heel-prick measuring immunoreactive (IRT) levels; elevated IRT prompts confirmatory via NGS to detect CFTR frameshifts among other mutations. These methods have significantly reduced morbidity by enabling timely interventions, though they focus on high-risk or screened conditions rather than genome-wide indel detection. Frameshift mutations underlie certain inherited disorders at frequencies around 1 in 10,000 births, depending on the condition and population; for instance, in , which has an overall incidence of about 1 in 3,000 live births, frameshifts represent a substantial subset of cases. In , somatic frameshift mutations are prevalent in approximately 15% of colorectal tumors exhibiting high , where they drive oncogenesis through loss-of-function in tumor suppressor genes. These estimates highlight the clinical relevance of screening in both and contexts. Distinguishing pathogenic frameshift mutations from benign poses significant diagnostic challenges, addressed through standardized frameworks like the ACMG/AMP guidelines. Null variants such as frameshifts in genes where loss-of-function is a known mechanism receive a "very strong" pathogenic score (PVS1), but requires integrating , computational predictions, and functional evidence to avoid misclassification, especially for novel in regions of variable tolerance. Variants of uncertain significance (VUS) are common in indel testing, necessitating periodic re-evaluation as databases grow, to ensure accurate in clinical settings.

Programmed Ribosomal Frameshifting

Mechanism in viruses and eukaryotes

Programmed ribosomal frameshifting (PRF) is a regulated translational recoding event in which the intentionally slips by one in the 5' (-1 PRF) or 3' (+1 PRF) direction during mRNA decoding, allowing the production of distinct protein isoforms from a single mRNA transcript. This contrasts with error-prone frameshifts caused by , as PRF is precisely controlled by cis-acting mRNA elements to ensure accurate alternative protein synthesis. In viruses, PRF is essential for expressing polyproteins that combine structural and enzymatic functions, optimizing compact genomes. A prominent example is human immunodeficiency virus type 1 (HIV-1), where -1 PRF at the gag-pol overlap site produces the Gag-Pol , incorporating , , and integrase domains necessary for . The efficiency of this -1 PRF in HIV-1 is approximately 5%, maintaining an optimal Gag-to-Gag-Pol ratio of about 20:1 for virion assembly, and is tightly regulated to prevent overproduction of enzymatic components. Similar -1 PRF mechanisms occur in other retroviruses and many positive-strand viruses, enabling the translation of replicase genes. Eukaryotic organisms also employ PRF, though less frequently than viruses, to regulate in cellular contexts. In (), +1 PRF is used in the EST3 gene, which encodes a component of the telomerase complex; frameshifting produces the full-length Est3p protein required for maintenance, with of 75-90%. Additionally, in retrotransposons like Ty3, PRF increases more than twofold during amino acid starvation, a stress response that enhances transposon activity. In , +1 PRF occurs in the ornithine decarboxylase antizyme (OAZ) gene, generating a full-length protein that regulates levels, a process linked to cellular stress adaptation; the frameshift is modulated by upstream open reading frames and concentrations. The core structural elements driving PRF include slippery heptanucleotide sequences in the mRNA, such as XXXYYYZ for -1 PRF (where X, Y, and Z represent specific allowing tRNA slippage, e.g., UUUUUUA in HIV-1), paired with downstream stimulatory RNA structures like pseudoknots or stem-loops that pause the and promote frameshifting. In +1 PRF cases, such as in and , motifs often involve P-site codon-anticodon pairing disruptions and upstream Shine-Dalgarno-like sequences, with pseudoknots or hairpins enhancing efficiency by stabilizing the paused ribosomal state. These elements ensure frameshifting occurs at specific sites with , typically without requiring factors in basal conditions.

Biological significance

Programmed ribosomal frameshifting (PRF) contributes significantly to gene economy in viruses by enabling the of multiple functional proteins from a single , thus maximizing coding capacity within constrained genome sizes. This mechanism is particularly vital for viruses, where genome compactness is essential for efficient replication and packaging. For instance, in coronaviruses like , a -1 PRF event at the junction of ORF1a and ORF1b allows ribosomes to shift frames and produce the ORF1ab polyprotein, which includes essential replicase components, thereby avoiding the need for separate promoters or initiation sites. Beyond structural efficiency, PRF exerts precise regulatory control over protein and adapts to environmental cues. In retroviruses such as HIV-1, -1 PRF at the gag-pol overlap site produces the Gag-Pol at a low frequency (approximately 1-5%), maintaining an optimal Gag:Pol ratio that balances viral assembly with enzymatic function while preventing excess toxicity. PRF efficiency can also be modulated by cellular stress or antiviral responses; factors like RNA-binding proteins or small molecules alter slippage rates, expression during or host activation.00044-4) From an evolutionary perspective, PRF elements exhibit remarkable conservation across viral taxa and select cellular systems, underscoring their adaptive value. This recoding strategy is widespread among positive-strand viruses, including retroviruses, coronaviruses, and flaviviruses, facilitating genome optimization over millions of years. In , the +1 PRF in the prfB , which encodes 2, traces back to the last common ancestor, with slippery sequences and stimulatory structures preserved to regulate termination efficiency. Although infrequent in eukaryotic genomes, PRF appears in specialized contexts, such as yeast prion-like domains that enhance -1 frameshifting for stress , and in bacterial genes where antibiotic-induced PRF, as seen with , drives expression of resistance determinants. Recent investigations (2023–2025) have illuminated PRF's implications in human pathophysiology, particularly neurodegeneration, where perturbed frameshifting disrupts protein homeostasis and generates aberrant isoforms. In repeat-expansion disorders like (ALS) and (FTD), altered PRF at expanded tracts leads to shifted reading frames, producing toxic repeats that exacerbate neuronal damage through imbalanced ratios of wild-type to mutant proteins. These findings highlight PRF's dual role in normal and disease, with frameshifting efficiency influenced by structures and cellular stressors in affected neurons.

Pathological Associations

Inherited genetic disorders

Frameshift mutations in the gene, which encodes the alpha subunit of beta-hexosaminidase A, are a primary cause of Tay-Sachs disease, a fatal lysosomal storage disorder characterized by progressive neurodegeneration due to accumulation of GM2 gangliosides. The most common such among Ashkenazi Jewish individuals is a 4-base pair insertion (1278insTATC) in 11, leading to a premature and complete loss of activity. This frameshift accounts for approximately 75-80% of disease-causing alleles in this population, where the carrier frequency is about 1 in 27, contributing to an incidence of 1 in 3,600 births. The resulting deficiency disrupts lipid degradation in neurons, leading to severe developmental regression, seizures, and death by early childhood. In , frameshift mutations in the CFTR gene disrupt the protein, impairing chloride ion transport and causing thick buildup in organs like the lungs and . These mutations, classified as class I defects, represent about 5% of all CFTR variants and typically result in no functional protein due to premature termination. An illustrative example is the 3905insT mutation (c.3773_3774insT) in 19, a 1-base pair insertion that causes a frameshift and premature , yielding a nonfunctional protein. This leads to classic symptoms including recurrent infections, pancreatic insufficiency, and reduced , with frameshifts contributing to severe disease phenotypes in affected individuals. Frameshift mutations in the gene (also known as CARD15) are strongly associated with , an inflammatory bowel disorder involving chronic intestinal due to dysregulated immune responses to . The 3020insC insertion in 11, one of three major NOD2 variants, causes a truncated protein that impairs recognition of bacterial peptidoglycans, leading to excessive and reduced autophagy.80138-X/fulltext) This frameshift is found in 10-30% of Crohn's patients, particularly those with ileal involvement, increasing disease risk 3- to 40-fold depending on and conferring susceptibility to structuring complications. The resulting immune dysregulation promotes Th1-mediated granulomatous , exacerbating tissue damage in the gastrointestinal tract.00169-X) Smith-Magenis syndrome, a , arises from frameshift in the RAI1 gene on chromosome 17p11.2, causing of the retinoic acid-inducible 1 protein, which regulates in neural and metabolic pathways. These , often small insertions or deletions leading to premature , account for about 10% of non-deletion cases and result in , sleep disturbances, and behavioral issues like self-injurious behavior. Affected individuals exhibit delayed speech, motor skills deficits, and autism-like features due to disrupted and synaptic function. A hotspot for such frameshifts exists in the polyglutamine repeat region of RAI1, amplifying the dosage-sensitive effects on brain development. Frameshift mutations in sarcomeric genes such as MYH7 (encoding beta-myosin heavy chain) and TNNT2 (encoding cardiac ) contribute to , a monogenic condition marked by abnormal thickening of the heart muscle and risk of arrhythmias or . In MYH7, frameshifts like c.5769delG produce truncated proteins that disrupt assembly and force generation in cardiomyocytes, leading to asymmetric septal . Similarly, frameshifts in TNNT2, such as those causing early termination, alter calcium sensitivity and function, promoting hypercontractility and . These variants, though less common than missense mutations, are linked to early-onset disease and sudden cardiac death in families, emphasizing the role of protein truncation in instability.

Role in oncogenesis

Frameshift mutations play a critical role in oncogenesis by introducing somatic alterations that disrupt tumor suppressor genes and drive tumorigenesis, particularly in cancers with defective mechanisms. These mutations often result in truncated or aberrant proteins that lose functional domains, thereby promoting uncontrolled and genomic instability. In , frameshift mutations in the gene are a hallmark initiating event, frequently occurring at hotspots like codon 1309, leading to the inactivation of this key regulator of the and facilitating formation. Similarly, in and ovarian cancers, frameshift indels in are prevalent, with over 1,800 distinct mutations reported in databases like , many of which cause premature termination and impair repair, elevating cancer risk. A major mechanism amplifying frameshift mutations in cancer is microsatellite instability (MSI) arising from mismatch repair (MMR) deficiency, which impairs the correction of replication errors in repetitive DNA sequences. MSI-high (MSI-H) tumors exhibit elevated rates of frameshift mutations, particularly in coding microsatellites of tumor suppressors and oncogenes, contributing to approximately 15% of all colorectal cancers. This deficiency leads to a hypermutated phenotype, with frameshifts enriched in MSI-H tumors across various cancer types, significantly increasing the tumor mutational burden (TMB) and fostering an environment conducive to tumor evolution and immune evasion. For instance, in MMR-deficient colorectal cancers, recurrent frameshifts in genes like TGFBR2 and ACVR2A disrupt TGF-β signaling, promoting metastatic progression. Beyond inactivation, frameshift mutations generate novel immunogenic neoantigens by producing out-of-frame peptides that can be presented on MHC molecules, potentially eliciting antitumor immune responses. These neoantigens are particularly abundant in MSI-H tumors due to their high frameshift burden, offering opportunities for . Recent studies have identified shared frameshift-derived neoantigens in (RCC), where personalized neoantigen targeting these mutations induced durable T-cell responses and prevented recurrence in high-risk patients, as demonstrated in a 2025 . Such findings highlight the dual oncogenic and therapeutic potential of frameshifts, with their frequency correlating inversely to immunogenicity in some contexts, guiding design strategies.

Implications in infectious diseases

Frameshift mutations play a significant role in host-pathogen interactions during infection. The Δ32 variant, a 32-base-pair deletion in the , introduces a premature via frameshift, resulting in a truncated, non-functional receptor that prevents entry into + T cells and confers near-complete resistance to R5-tropic -1 strains in homozygous individuals. This mutation, prevalent in about 10-15% of populations, highlights how frameshifts can evolve as protective adaptations against viral invasion. In HIV itself, programmed -1 ribosomal frameshifting is essential for expressing the Pol polyprotein from the gag-pol overlapping reading frame, maintaining a precise Gag-to-Gag-Pol ratio of approximately 20:1 for viral assembly and replication. However, spontaneous or induced frameshift mutations that disrupt this signal—such as alterations in the slippery sequence or stimulatory stem-loop—reduce frameshifting efficiency by 30-60%, leading to imbalanced protein production and diminished viral replication fitness in cell culture and animal models. These fitness costs underscore the evolutionary pressure to preserve programmed frameshifting in HIV quasispecies. Bacterial pathogens also exploit frameshift mutations for adaptation, particularly in acquiring antibiotic resistance. For instance, frameshifts in the rpoB gene, encoding the β subunit, can suppress deleterious effects of rifampicin resistance mutations, enabling high-level resistance without severe fitness penalties through intrinsic translational suppression. Similarly, frameshift indels in regulatory genes like mmpR5 in species disrupt repressor function, upregulating efflux pumps and conferring resistance to , a key drug for . Such mutations illustrate how frameshifts facilitate rapid evolutionary responses to selective pressures in bacterial populations. In RNA viruses like A, mutational frameshifts contribute to quasispecies diversity, enabling to immune pressures and environmental changes. High error-prone replication generates frequent , and compensatory frameshifts—where a second indel restores the —allow preservation of functional protein domains while introducing sequence variation, as observed in hundreds of influenza genes across global isolates. This mechanism drives intra- evolution and antigenic drift, enhancing viral persistence and evasion of . Host antiviral defenses further implicate frameshifts in infectious disease dynamics through APOBEC3-mediated hypermutation of viral genomes. APOBEC3G, a deaminase, induces extensive G-to-A in -1 reverse transcripts, preferentially targeting GG dinucleotides to generate premature stop codons (e.g., TGG to TAG), which truncate viral proteins in a manner akin to frameshift-induced disruption, often rendering progeny virions non-infectious. This hypermutation strategy, evaded by HIV Vif but potent in its absence, exemplifies how host-induced genetic alterations limit viral propagation across diverse pathogens.

Therapeutic Strategies

Corrective gene editing

Corrective gene editing employs precision genome engineering technologies to directly repair or reverse frameshift mutations, restoring the proper and protein function in affected . Among these, CRISPR-Cas9 systems facilitate correction through (HDR), where a donor template guides the precise insertion or deletion of to counteract the frameshift caused by non-multiples-of-three indels. This approach has demonstrated up to 30-fold enhancement in HDR efficiency for repairing frameshift mutations in cellular assays, enabling restoration of function without reliance on error-prone (NHEJ). For (CFTR) mutations, preclinical studies from 2023 to 2025 explore CRISPR-Cas9 variants, including base editing strategies that introduce precise changes without double-strand breaks (DSBs) to mitigate frameshift effects, such as by correcting associated splicing defects or revertant sequences in patient-derived airway cells (as of November 2025). Prime editing represents a more versatile DSB-free method for frameshift reversal, utilizing a fusion of nickase, , and a prime editing (pegRNA) to directly install small insertions or deletions at the mutation site. This technique achieves correction efficiencies of 20-50% in human cell models for various frameshift-inducing indels, surpassing traditional rates in non-dividing cells and minimizing unintended genomic alterations. In applications targeting CFTR mutations, has restored function in primary airway epithelial organoids from patients, though primarily demonstrated for point mutations like L227R and N1303K, with optimized systems yielding up to 25% editing rates (as of 2024). Representative examples illustrate the therapeutic potential of these tools. In Tay-Sachs disease models, CRISPR-based editing of the gene corrects frameshift mutations by base editing or in neuronal cells, partially restoring β-hexosaminidase A activity and reducing accumulation. Similarly, studies on von Hippel-Lindau (VHL) frameshifts in employ to model and investigate neoantigen generation, where editing introduces or corrects frameshift indels to enhance tumor-specific immune responses, informing personalized strategies against driver mutations prevalent in over 90% of clear cell renal cancers. Despite these advances, challenges persist, including off-target effects from unintended cleavage and inefficient delivery to target tissues like the lungs or brain. Recent innovations, such as enhanced specificity variants and nanoparticle carriers, have improved editing precision by reducing off-target rates to below 1% in 2025 preclinical reviews, while addressing delivery hurdles through tissue-specific (as of 2025). As of 2025, base editing variants have shown promise in preclinical models for precise frameshift correction without DSBs, though clinical translation remains challenged by delivery and no FDA-approved gene therapies exist for frameshift disorders. These developments underscore the shift toward safer, more efficient corrective strategies for frameshift-associated disorders like and inherited neuropathies.

Pharmacological and other interventions

Pharmacological interventions for frameshift mutations primarily target the downstream consequences of these genetic alterations, such as premature termination codons (PTCs) that arise from reading frame disruptions, leading to truncated or nonfunctional proteins. Read-through drugs, particularly aminoglycosides like gentamicin, promote ribosomal of PTCs primarily induced by mutations, allowing translation to continue and produce full-length proteins; however, for frameshifts, efficacy is limited as read-through does not restore the original , resulting in aberrant proteins. In (CF), class I mutations in the CFTR gene (primarily , but including some frameshifts leading to PTCs and absent CFTR protein) have been investigated with gentamicin, showing partial restoration in patient-derived cells, though clinical benefits are modest due to toxicity and limited applicability to frameshifts. Similarly, in (DMD), mutations in the gene are targeted by gentamicin and other aminoglycosides such as and , demonstrating suppression of stops and increased dystrophin in preclinical models; for frameshift mutations (out-of-frame deletions), is the preferred approach over read-through. These agents bind to the ribosomal decoding site, stabilizing near-cognate tRNAs at PTCs, with efficacy enhanced by a uracil upstream of the . Chaperone therapies aim to stabilize misfolded proteins resulting from certain mutations, but for frameshifts causing PTCs, little protein is produced for correction unless partial occurs. In , chemical chaperones such as and 4-phenylbutyrate have been investigated to correct folding defects in mutant CFTR proteins, enhancing trafficking to the and activity in cellular models. Pharmacological chaperones like lumacaftor (VX-809), approved for , primarily target class II mutations such as the common ΔF508 in-frame deletion and have shown investigational potential in combination with read-through agents for class I mutations by stabilizing rescued CFTR, though specificity for frameshift-derived proteins remains limited. Clinical studies indicate modest improvements in function when chaperones are used adjunctively, though primarily for non-frameshift variants. Symptom management strategies for frameshift mutation-associated disorders focus on mitigating disease effects rather than correcting the genetic lesion. In Tay-Sachs disease, caused by frameshift mutations in the HEXA gene leading to hexosaminidase A deficiency and GM2 ganglioside accumulation, enzyme replacement therapy (ERT) delivers recombinant hexosaminidase A to lysosomes, reducing substrate buildup in preclinical and early clinical settings. For instance, intrathecal ERT has improved neurological outcomes in animal models of late-onset Tay-Sachs by crossing the blood-brain barrier. In oncology, frameshift mutations in microsatellite instability-high (MSI-H) cancers generate immunogenic neoantigens due to altered peptide sequences; immunotherapy with immune checkpoint inhibitors like pembrolizumab exploits these neoantigens, with tumor frameshift burden predicting response rates up to 50% in MSI-H colorectal cancers. Shared frameshift-derived neoantigens, such as those from recurrent indels in TGFB2R and ACVR2A, enable off-the-shelf vaccines that elicit CD8+ T cell responses in multiple patients. Emerging interventions include designed to skip exons harboring frameshift mutations, restoring the and producing partially functional proteins. In DMD, ASOs like target exons disrupted by frameshifts, inducing skipping to yield in-frame transcripts and increase dystrophin levels by 5-10% in muscle biopsies of treated patients. Cocktails of ASOs have shown promise for multi-exon skips in frameshift cases, with phase III trials demonstrating improved ambulation. For antiviral applications, modulators of programmed ribosomal frameshifting (PRF) inhibit by disrupting -1 PRF signals in coronaviruses. Recent 2024-2025 studies identified small-molecule inhibitors from high-throughput screens that reduce PRF efficiency by over 70%, impeding replication in cell cultures without host toxicity, paving the way for broad-spectrum antivirals targeting frameshift-dependent viral polyproteins. A 2025 detailed novel frameshift inhibitors derived from natural products, effective against multiple coronaviruses at micromolar concentrations.