Editing Alternative splicing (section)

==Disease==
Changes in the RNA processing machinery may lead to mis-splicing of multiple transcripts, while single-nucleotide alterations in splice sites or cis-acting splicing regulatory sites may lead to differences in splicing of a single gene, and thus in the mRNA produced from a mutant gene's transcripts. A study in 2005 involving probabilistic analyses indicated that greater than 60% of human disease-causing [[mutation]]s affect splicing rather than directly affecting coding sequences.<ref name="Lopez-Bigas">{{cite journal | vauthors = López-Bigas N, Audit B, Ouzounis C, Parra G, Guigó R | title = Are splicing mutations the most frequent cause of hereditary disease? | journal = FEBS Letters | volume = 579 | issue = 9 | pages = 1900–3 | date = March 2005 | pmid = 15792793 | doi = 10.1016/j.febslet.2005.02.047 | s2cid = 30174458 | doi-access =  | bibcode = 2005FEBSL.579.1900L }}</ref> A more recent study indicates that one-third of all hereditary diseases are likely to have a splicing component.<ref name="Lim" /> Regardless of exact percentage, a number of splicing-related diseases do exist.<ref>{{cite journal | vauthors = Ward AJ, Cooper TA | title = The pathobiology of splicing | journal = The Journal of Pathology | volume = 220 | issue = 2 | pages = 152–63 | date = January 2010 | pmid = 19918805 | pmc = 2855871 | doi = 10.1002/path.2649 }}</ref> As described below, a prominent example of splicing-related diseases is cancer.

Abnormally spliced mRNAs are also found in a high proportion of cancerous cells.<ref name="Skotheim and Nees 2007">{{cite journal | vauthors = Skotheim RI, Nees M | title = Alternative splicing in cancer: noise, functional, or systematic? | journal = The International Journal of Biochemistry & Cell Biology | volume = 39 | issue = 7–8 | pages = 1432–49 | year = 2007 | pmid = 17416541 | doi = 10.1016/j.biocel.2007.02.016 }}</ref><ref name="He2009">{{cite journal | vauthors = He C, Zhou F, Zuo Z, Cheng H, Zhou R | title = A global view of cancer-specific transcript variants by subtractive transcriptome-wide analysis | journal = PLOS ONE | volume = 4 | issue = 3 | pages = e4732 | year = 2009 | pmid = 19266097 | pmc = 2648985 | doi = 10.1371/journal.pone.0004732 | veditors = Bauer JA | bibcode = 2009PLoSO...4.4732H | doi-access = free }}</ref><ref name = "Sveen 2015">{{cite journal | vauthors = Sveen A, Kilpinen S, Ruusulehto A, Lothe RA, Skotheim RI | title = Aberrant RNA splicing in cancer; expression changes and driver mutations of splicing factor genes | journal = Oncogene | volume = 35 | issue = 19 | pages = 2413–27 | date = May 2016 | pmid = 26300000 | doi = 10.1038/onc.2015.318 | s2cid = 22943729 | doi-access = free }}</ref> Combined [[RNA-Seq]] and proteomics analyses have revealed striking differential expression
of splice isoforms of key proteins in important cancer pathways.<ref>{{cite journal | vauthors = Omenn GS, Guan Y, Menon R | title = A new class of protein cancer biomarker candidates: differentially expressed splice variants of ERBB2 (HER2/neu) and ERBB1 (EGFR) in breast cancer cell lines | journal = Journal of Proteomics | volume = 107 | pages = 103–12 | date = July 2014 | pmid = 24802673 | pmc = 4123867 | doi = 10.1016/j.jprot.2014.04.012 }}</ref> It is not always clear whether such aberrant patterns of splicing contribute to the cancerous growth, or are merely consequence of cellular abnormalities associated with cancer. For certain types of cancer, like in colorectal and prostate, the number of splicing errors per cancer has been shown to vary greatly between individual cancers, a phenomenon referred to as [[transcriptome instability]].<ref name="Sveen 2011">{{cite journal | vauthors = Sveen A, Johannessen B, Teixeira MR, Lothe RA, Skotheim RI | title = Transcriptome instability as a molecular pan-cancer characteristic of carcinomas | journal = BMC Genomics | volume = 15 | pages = 672 | date = August 2014 | issue = 1 | pmid = 25109687 | pmc = 3219073 | doi = 10.1186/1471-2164-15-672 | doi-access = free }}</ref><ref name="Sveen 2014">{{cite journal | vauthors = Sveen A, Agesen TH, Nesbakken A, Rognum TO, Lothe RA, Skotheim RI | title = Transcriptome instability in colorectal cancer identified by exon microarray analyses: Associations with splicing factor expression levels and patient survival | journal = Genome Medicine | volume = 3 | issue = 5 | pages = 32 | date = May 2011 | pmid = 21619627 | pmc = 4137096 | doi = 10.1186/gm248 | doi-access = free }}</ref> Transcriptome instability has further been shown to correlate grealty with reduced expression level of splicing factor genes. Mutation of ''[[DNMT3A]]'' has been demonstrated to contribute to [[hematologic malignancies]], and that ''[[DNMT3A]]''-mutated cell lines exhibit [[transcriptome instability]] as compared to their isogenic<!-- That link is a disambiguation to [[Zygosity]], which doesn't even contain the term, and [[Isogenic human disease models]], which is specifically NOT wildtype. --> wildtype counterparts.<ref name="Banaszak 2018">{{cite journal | vauthors = Banaszak LG, Giudice V, Zhao X, Wu Z, Gao S, Hosokawa K, Keyvanfar K, Townsley DM, Gutierrez-Rodrigues F, Fernandez Ibanez MD, Kajigaya S, Young NS | display-authors = 6 | title = Abnormal RNA splicing and genomic instability after induction of DNMT3A mutations by CRISPR/Cas9 gene editing | journal = Blood Cells, Molecules & Diseases | volume = 69 | pages = 10–22 | date = March 2018 | pmid = 29324392 | pmc = 6728079 | doi = 10.1016/j.bcmd.2017.12.002 }}</ref>

In fact, there is actually a reduction of alternative splicing in cancerous cells compared to normal ones, and the types of splicing differ; for instance, cancerous cells show higher levels of intron retention than normal cells, but lower levels of exon skipping.<ref name="Kim2008">{{cite journal | vauthors = Kim E, Goren A, Ast G | title = Insights into the connection between cancer and alternative splicing | journal = Trends in Genetics | volume = 24 | issue = 1 | pages = 7–10 | date = January 2008 | pmid = 18054115 | doi = 10.1016/j.tig.2007.10.001 }}</ref> Some of the differences in splicing in cancerous cells may be due to the high frequency of somatic mutations in splicing factor genes,<ref name="Sveen 2015"/> and some may result from changes in [[phosphorylation]] of trans-acting splicing factors.<ref name="Fackenthal">{{cite journal | vauthors = Fackenthal JD, Godley LA | title = Aberrant RNA splicing and its functional consequences in cancer cells | journal = Disease Models & Mechanisms | volume = 1 | issue = 1 | pages = 37–42 | year = 2008 | pmid = 19048051 | pmc = 2561970 | doi = 10.1242/dmm.000331 | format = Free full text }}</ref> Others may be produced by changes in the relative amounts of splicing factors produced; for instance, breast cancer cells have been shown to have increased levels of the splicing factor [[SF2/ASF]].<ref name="Ghigna2005">{{cite journal | vauthors = Ghigna C, Giordano S, Shen H, Benvenuto F, Castiglioni F, Comoglio PM, Green MR, Riva S, Biamonti G | display-authors = 6 | title = Cell motility is controlled by SF2/ASF through alternative splicing of the Ron protooncogene | journal = Molecular Cell | volume = 20 | issue = 6 | pages = 881–90 | date = December 2005 | pmid = 16364913 | doi = 10.1016/j.molcel.2005.10.026 | doi-access = free }}</ref> One study found that a relatively small percentage (383 out of over 26000) of alternative splicing variants were significantly higher in frequency in tumor cells than normal cells, suggesting that there is a limited set of genes which, when mis-spliced, contribute to tumor development.<ref name="pmid15048092">{{cite journal | vauthors = Hui L, Zhang X, Wu X, Lin Z, Wang Q, Li Y, Hu G | title = Identification of alternatively spliced mRNA variants related to cancers by genome-wide ESTs alignment | journal = Oncogene | volume = 23 | issue = 17 | pages = 3013–23 | date = April 2004 | pmid = 15048092 | doi = 10.1038/sj.onc.1207362 | doi-access = free }}</ref> It is believed however that the deleterious effects of mis-spliced transcripts are usually safeguarded and eliminated by a cellular posttranscriptional quality control mechanism termed [[nonsense-mediated mRNA decay]] [NMD].<ref name="Danckwardt2002">{{cite journal | vauthors = Danckwardt S, Neu-Yilik G, Thermann R, Frede U, Hentze MW, Kulozik AE | title = Abnormally spliced beta-globin mRNAs: a single point mutation generates transcripts sensitive and insensitive to nonsense-mediated mRNA decay | journal = Blood | volume = 99 | issue = 5 | pages = 1811–6 | date = March 2002 | pmid = 11861299 | doi = 10.1182/blood.V99.5.1811 | s2cid = 17128174 | doi-access = free }}</ref>

One example of a specific splicing variant associated with cancers is in one of the human [[DNMT]] genes. Three DNMT genes encode enzymes that add [[DNA methylation|methyl]] groups to DNA, a modification that often has regulatory effects. Several abnormally spliced DNMT3B mRNAs are found in tumors and cancer cell lines. In two separate studies, expression of two of these abnormally spliced mRNAs in mammalian cells caused changes in the DNA methylation patterns in those cells. Cells with one of the abnormal mRNAs also grew twice as fast as control cells, indicating a direct contribution to tumor development by this product.<ref name="Fackenthal" />

Another example is the ''Ron'' (''[[MST1R]]'') [[proto-oncogene]]. An important property of cancerous cells is their ability to move and invade normal tissue. Production of an abnormally spliced transcript of ''Ron'' has been found to be associated with increased levels of the SF2/ASF in breast cancer cells. The abnormal isoform of the Ron protein encoded by this mRNA leads to [[cell motility]].<ref name="Ghigna2005" />

Overexpression of a truncated splice variant of the [[FOSB]] gene – [[ΔFosB]] – in a specific population of neurons in the [[nucleus accumbens]] has been identified as the causal mechanism involved in the induction and maintenance of an [[addiction]] to drugs and [[natural reward]]s.<ref name="Cellular basis">{{cite journal | vauthors = Nestler EJ | title = Cellular basis of memory for addiction | journal = Dialogues in Clinical Neuroscience | volume = 15 | issue = 4 | pages = 431–43 | date = December 2013 | pmid = 24459410 | pmc = 3898681 | doi = 10.31887/DCNS.2013.15.4/enestler | quote = DESPITE THE IMPORTANCE OF NUMEROUS PSYCHOSOCIAL FACTORS, AT ITS CORE, DRUG ADDICTION INVOLVES A BIOLOGICAL PROCESS: the ability of repeated exposure to a drug of abuse to induce changes in a vulnerable brain that drive the compulsive seeking and taking of drugs, and loss of control over drug use, that define a state of addiction.&nbsp;... A large body of literature has demonstrated that such ΔFosB induction in D1-type NAc neurons increases an animal's sensitivity to drug as well as natural rewards and promotes drug self-administration, presumably through a process of positive reinforcement }}</ref><ref name="What the ΔFosB?">{{cite journal | vauthors = Ruffle JK | title = Molecular neurobiology of addiction: what's all the (Δ)FosB about? | journal = The American Journal of Drug and Alcohol Abuse | volume = 40 | issue = 6 | pages = 428–37 | date = November 2014 | pmid = 25083822 | doi = 10.3109/00952990.2014.933840 | quote = ΔFosB is an essential transcription factor implicated in the molecular and behavioral pathways of addiction following repeated drug exposure. The formation of ΔFosB in multiple brain regions, and the molecular pathway leading to the formation of AP-1 complexes is well understood. The establishment of a functional purpose for ΔFosB has allowed further determination as to some of the key aspects of its molecular cascades, involving effectors such as GluR2 (87,88), Cdk5 (93) and NFkB (100). Moreover, many of these molecular changes identified are now directly linked to the structural, physiological and behavioral changes observed following chronic drug exposure (60,95,97,102). New frontiers of research investigating the molecular roles of ΔFosB have been opened by epigenetic studies, and recent advances have illustrated the role of ΔFosB acting on DNA and histones, truly as a ''molecular switch'' (34). As a consequence of our improved understanding of ΔFosB in addiction, it is possible to evaluate the addictive potential of current medications (119), as well as use it as a biomarker for assessing the efficacy of therapeutic interventions (121,122,124). Some of these proposed interventions have limitations (125) or are in their infancy (75). However, it is hoped that some of these preliminary findings may lead to innovative treatments, which are much needed in addiction. | s2cid = 19157711 }}</ref><ref name="G9a reverses ΔFosB plasticity">{{cite journal | vauthors = Biliński P, Wojtyła A, Kapka-Skrzypczak L, Chwedorowicz R, Cyranka M, Studziński T | title = Epigenetic regulation in drug addiction | journal = Annals of Agricultural and Environmental Medicine | volume = 19 | issue = 3 | pages = 491–6 | year = 2012 | pmid = 23020045 | quote = For these reasons, ΔFosB is considered a primary and causative transcription factor in creating new neural connections in the reward centre, prefrontal cortex, and other regions of the limbic system. This is reflected in the increased, stable and long-lasting level of sensitivity to cocaine and other drugs, and tendency to relapse even after long periods of abstinence. These newly constructed networks function very efficiently via new pathways as soon as drugs of abuse are further taken }}</ref><ref name="Natural and drug addictions">{{cite journal | vauthors = Olsen CM | title = Natural rewards, neuroplasticity, and non-drug addictions | journal = Neuropharmacology | volume = 61 | issue = 7 | pages = 1109–22 | date = December 2011 | pmid = 21459101 | pmc = 3139704 | doi = 10.1016/j.neuropharm.2011.03.010 }}</ref>

Recent provocative studies point to a key function of chromatin structure and histone modifications in alternative splicing regulation. These insights suggest that epigenetic regulation determines not only what parts of the genome are expressed but also how they are spliced.<ref name="Luco2011">{{cite journal | vauthors = Luco RF, Allo M, Schor IE, Kornblihtt AR, Misteli T | title = Epigenetics in alternative pre-mRNA splicing | journal = Cell | volume = 144 | issue = 1 | pages = 16–26 | date = January 2011 | pmid = 21215366 | pmc = 3038581 | doi = 10.1016/j.cell.2010.11.056 }}</ref>