Editing Frameshift mutation (section)

==Mechanism==

Frameshift mutations can occur randomly or be caused by an external stimulus. The detection of frameshift mutations can occur via several different methods. Frameshifts are just one type of mutation that can lead to incomplete or incorrect proteins, but they account for a significant percentage of errors in DNA.In an unaltered gene, codons (triplets of nucleotides) are sequentially interpreted, with each codon encoding a specific amino acid. This is known as the standard reading frame. However, in cases of frame shift mutations, an extra nucleotide (or more) is inserted into the DNA sequence, disrupting the typical reading frame and causing a shift in the sequence.

This insertion prompts a shift in the reading frame due to the triplet nature of the genetic code. For instance, the addition of an extra "A" leads to a sequence shift, triggering the reading of an entirely different set of codons. This deviation in genetic information causes the ribosome, which reads the mRNA for protein synthesis, to misinterpret the genetic data. Consequently, an entirely different series of amino acids is generated, resulting in the generation of an altered protein sequence. In most instances, the new reading frame results in an early encounter with a stop codon, leading to the formation of a shortened and usually inactive protein. This form of mutation is termed an early stop codon or a nonsense mutation.

===Genetic or environmental===

{{Main article|Mutation}}

This is a genetic mutation at the level of nucleotide bases. Why and how frameshift mutations occur are continually being sought after. An environmental study, specifically the production of [[UV]]-induced frameshift mutations by DNA polymerases deficient in 3′ → 5′ exonuclease activity was done. The normal sequence 5′ GTC GTT TTA CAA 3′ was changed to GTC GTT T TTA CAA (MIDT) of GTC GTT C TTA CAA (MIDC) to study frameshifts. [[E. coli]] pol I Kf and T7 DNA polymerase mutant [[enzymes]] devoid of 3′ → 5′ exonuclease activity produce UV-induced revertants at higher frequency than did their [[exonuclease]] proficient counterparts. The data indicates that loss of proofreading activity increases the frequency of UV-induced frameshifts.<ref>{{cite journal|last=Sagher|first=Daphna|author2=Turkington, Edith|author3= Acharya, Sonia|author4= Strauss, Bernard|title=Production of UV-induced Frameshift Mutations in Vitro by DNA Polymerases Deficient in 3′ → 5′ Exonuclease Activity|journal=Journal of Molecular Biology|volume=240|issue=3|pages=226–242|doi=10.1006/jmbi.1994.1437 |pmid=8028006|date=July 1994}}</ref>

===Detection===

====Fluorescence====

The effects of neighboring bases and secondary structure to detect the frequency of frameshift mutations has been investigated in depth using [[fluorescence]]. Fluorescently tagged DNA, by means of base analogues, permits one to study the local changes of a DNA sequence.<ref>{{cite journal |first=Neil P. |last=Johnson |author2=Walter A. Baase |author3=Peter H. von Hippel  |title=Low-energy circular dichroism of 2-aminopurine dinucleotide as a probe of local conformation of DNA and RNA |journal=Proc Natl Acad Sci U S A |date=March 2004 |volume=101 |issue=10 |pages=3426–31 |doi=10.1073/pnas.0400591101 |pmid=14993592 |pmc=373478|bibcode=2004PNAS..101.3426J |doi-access=free }}</ref> Studies on the effects of the length of the primer strand reveal that an equilibrium mixture of four hybridization conformations was observed when template bases looped-out as a bulge, i.e. a structure flanked on both sides by duplex DNA. In contrast, a double-loop structure with an unusual unstacked DNA conformation at its downstream edge was observed when the extruded bases were positioned at the primer–template junction, showing that misalignments can be modified by neighboring DNA secondary structure.<ref>{{cite journal |first=Walter A. |last=Baase |author2=Davis Jose  |author3=Benjamin C. Ponedel  |author4=Peter H. von Hippel  |author5=Neil P. Johnson  |title=DNA models of trinucleotide frameshift deletions: the formation of loops and bulges at the primer–template junction |journal=Nucleic Acids Research |doi=10.1093/nar/gkn1042 |volume=37 |issue=5 |pages=1682–9 |pmid=19155277 |pmc=2655659 |url=|year=2009 }}</ref>

====Sequencing====
[[File:Frameshift deletion (13062713935).jpg|thumb|A deletion mutation alters every codon following it, and can make protein synthesis stop prematurely by forming a [[stop codon]].]]
[[Sanger sequencing]] and [[pyrosequencing]] are two methods that have been used to detect frameshift mutations, however, it is likely that data generated will not be of the highest quality. Even still, 1.96 million [[indel]]s have been identified through Sanger sequencing that do not overlap with other databases. When a frameshift mutation is observed it is compared against the Human Genome Mutation Database (HGMD) to determine if the mutation has a damaging effect. This is done by looking at four features. First, the ratio between the affected and conserved DNA, second the location of the mutation relative to the transcript, third the ratio of conserved and affected amino acids and finally the distance of the indel to the end of the [[exon]].<ref name="predicting frameshifts" />

[[Massively parallel sequencing|Massively Parallel Sequencing]] is a newer method that can be used to detect mutations. Using this method, up to 17 gigabases can be sequenced at once, as opposed to limited ranges for [[Sanger sequencing]] of only about 1 kilobase. Several technologies are available to perform this test and it is being looked at to be used in clinical applications.<ref name="TuckerMarra2009">{{cite journal|last1=Tucker|first1=Tracy|last2=Marra|first2=Marco|last3=Friedman|first3=Jan M.|title=Massively Parallel Sequencing: The Next Big Thing in Genetic Medicine|journal=The American Journal of Human Genetics|volume=85|issue=2|year=2009|pages=142–154 |doi=10.1016/j.ajhg.2009.06.022|pmid=19679224|pmc=2725244}}</ref> When testing for different carcinomas, current methods only allow for looking at one gene at a time. Massively Parallel Sequencing can test for a variety of cancer causing mutations at once as opposed to several specific tests.<ref name="WalshCasadei2011">{{cite journal|last1 = Walsh|first1 = T.|last2 = Casadei|first2 = S.|last3 = Lee|first3 = M. K.|last4 = Pennil|first4 = C. C.|last5 = Nord|first5 = A. S.|last6 = Thornton|first6 = A. M.|last7 = Roeb|first7 = W.|last8 = Agnew|first8 = K. J.|last9 = Stray|first9 = S. M.|last10 = Wickramanayake|first10 = A.|last11 = Norquist|first11 = B.|last12 = Pennington|first12 = K. P.|last13 = Garcia|first13 = R. L.|last14 = King|first14 = M.-C.|last15 = Swisher|first15 = E. M.|title = From the Cover: Mutations in 12 genes for inherited ovarian, fallopian tube, and peritoneal carcinoma identified by massively parallel sequencing|journal =Proc Natl Acad Sci U S A |volume = 108|issue = 44|year = 2011|pages = 18032–7|doi = 10.1073/pnas.1115052108 |pmid=22006311 |pmc=3207658 |bibcode = 2011PNAS..10818032W|doi-access = free}}</ref> An experiment to determine the accuracy of this newer sequencing method tested for 21 genes and had no false positive calls for frameshift mutations.<ref name="WalshLee2010">{{cite journal|last1=Walsh|first1=T.|last2=Lee|first2=M. K.|last3=Casadei|first3=S.|last4=Thornton|first4=A. M.|last5=Stray|first5=S. M.|last6=Pennil|first6=C.|last7=Nord|first7=A. S.|last8=Mandell|first8=J. B.|last9=Swisher|first9=E. M.|last10=King|first10=M.-C.|title=Detection of inherited mutations for breast and ovarian cancer using genomic capture and massively parallel sequencing|journal=Proc Natl Acad Sci U S A |volume=107|issue=28|year=2010|pages=12629–33 |doi=10.1073/pnas.1007983107 |pmid=20616022 |pmc=2906584 |bibcode=2010PNAS..10712629W|doi-access=free}}</ref>

====Diagnosis====

A US [[patent]] (5,958,684) in 1999 by Leeuwen, details the methods and reagents for diagnosis of diseases caused by or associated with a gene having a somatic mutation giving rise to a frameshift mutation. The methods include providing a tissue or fluid sample and conducting gene analysis for frameshift mutation or a protein from this type of mutation. The nucleotide sequence of the suspected gene is provided from published gene sequences or from [[cloning]] and sequencing of the suspect gene. The amino acid sequence encoded by the gene is then predicted.<ref>US Patent [https://patents.google.com/patent/US5958684 5,958,684] (September 28, 1999) "Diagnosis of Neurodegenerative Disease" by Leeuwen ''et al''</ref> NA Sequencing: Sanger sequencing or Next-Generation Sequencing (NGS) can be used to directly sequence the DNA and identify insertions or deletions.Polymerase Chain Reaction (PCR): PCR can be used to amplify the specific region containing the mutation for subsequent analysis.Multiplex Ligation-dependent Probe Amplification (MLPA): MLPA is a technique used to detect copy number variations and small insertions or deletions.Comparative Genomic Hybridization (CGH): CGH is used to detect chromosomal imbalances, which may include large insertions or deletions.

====Frequency====

Despite the rules that govern the genetic code and the various mechanisms present in a cell to ensure the correct transfer of genetic information during the process of DNA replication as well as during translation, mutations do occur; frameshift mutation is not the only type. There are at least two other types of recognized point mutations, specifically [[missense mutation]] and [[nonsense mutation]].<ref name=MBoG_6th_2008/> A frameshift mutation can drastically change the coding capacity (genetic information) of the message.<ref name=MBoG_6th_2008/> Small insertions or deletions (those less than 20 base pairs) make up 24% of mutations that manifest in currently recognized genetic disease.<ref name="predicting frameshifts">{{cite journal|last=Hu|first=J|author2=Ng, PC|title=Predicting the effects of frameshifting indels.|journal=Genome Biology|date=9 February 2012|volume=13|issue=2|pages=R9|pmid=22322200 |doi=10.1186/gb-2012-13-2-r9|pmc=3334572|doi-access=free}}</ref>

Frameshift mutations are found to be more common in repeat regions of DNA. A reason for this is because of slipping of the polymerase enzyme in repeat regions, allowing for mutations to enter the [[sequence]].<ref name="editing frameshift mutation">{{cite journal|last=Harfe|first=BD|author2=Jinks-Robertson, S|title=Removal of frameshift intermediates by mismatch repair proteins in Saccharomyces cerevisiae.|journal=Molecular and Cellular Biology|date=July 1999|volume=19|issue=7|pages=4766–73|pmid=10373526|pmc=84275|doi=10.1128/MCB.19.7.4766}}</ref> [[Experiment]]s can be run to determine the frequency of the frameshift mutation by adding or removing a pre-set number of nucleotides. Experiments have been run by adding four basepairs, called the +4 experiments, but a team from [[Emory University]] looked at the difference in frequency of the mutation by both adding and deleting a base pair. It was shown that there was no difference in the frequency between the addition and deletion of a base pair. There is however, a difference in the result of the protein.<ref name="editing frameshift mutation" />

[[Huntington's disease]] is one of the nine codon reiteration disorders caused by polyglutamine expansion mutations that include spino-cerebellar ataxia (SCA) 1, 2, 6, 7 and 3, spinobulbar muscular atrophy and dentatorubal-pallidoluysianatrophy. There may be a link between diseases caused by polyglutamine and polyalanine expansion mutations, as frame shifting of the original SCA3 gene product encoding CAG/polyglutamines to GCA/polyalanines. Ribosomal slippage during translation of the SCA3 protein has been proposed as the mechanism resulting in shifting from the polyglutamine to the polyalanine-encoding frame. A dinucleotide deletion or single nucleotide insertion within the polyglutamine tract of huntingtin exon 1 would shift the CAG, polyglutamineen coding frame by +1 (+1 frame shift) to the GCA, polyalanine-encoding frame and introduce a novel epitope to the C terminus of Htt exon 1 (APAAAPAATRPGCG).<ref>{{cite journal|last=Davies|first=J E|author2=Rubinsztein, D C|title=Polyalanine and polyserine frameshift products in Huntington's disease|journal=Journal of Medical Genetics|volume=43|issue=11|pages=893–896|doi=10.1136/jmg.2006.044222 |pmid=16801344 |pmc=2563184 |year=2006}}</ref>