User:Jgould1400/sandbox

Week 3 Assignment: Summary of Five Pillars

1. As an encyclopedia, Wikipedia has elements of general and specialized encyclopedias but is not meant as a “soapbox”, “advertising platform”, or other type of “mouthpiece” for specific causes.

2. Wikipedia is written from a neutral point of view, using articles that document and explain major points of view in a balanced and impartial manner, and avoiding advocacy. We strive for verifiable accuracy and remove unreferenced material.

3. Wikipedia is free content that anyone can edit, use, modify, and distribute, but we respect copyright laws and do not plagiarize.

4. Editors interact with each other in a respectful and civil manner even if they disagree.

5. Wikipedia does not have firm rules: Be bold but not reckless in updating articles and don't worry about making mistakes.

Week 4 Assignment: Summary of Characteristics of Target Article

The Wikipedia editing goal for our Spring Semester 2013 Molecular Biology class will be to take an article from "Stub Class" to a level of quality somewhere between “B” level and "GA” level on Wikipedia's scale. “GA" stands for "Good Article” and is one level higher than "B" level.

A “GA” article is useful to nearly all readers and has no obvious problems. A “GA” article has a high level of citations to reliable sources, including good in-line citations; and the quality of a “GA” article is high enough that it approaches the quality of a professional encyclopedia: The information should be verifiable -- without the need to carry out additional original research.

A “B” level article is mostly complete and has no major issues. It should “sound right”, have good structure and “flow”, and have a suitable level of citations to reliable sources. A "B" article reasonably covers the topic and has no obvious omissions or inaccuracies. However, a "B" article still requires further work to reach “GA” level. For instance, the content may be helpful to most readers but may not be complete enough to satisfy serious researchers (expert help may be needed). In addition, the style of a "B" article may need some improvement and may need to be checked for compliance with reference to standard style guidelines.

Week 5 Assignment: Article Review

Glick et al., Spring Harb Perspect Biol 2011;3:a005215

In their 2011 article regarding Golgi Traffic, Glick BS, Luini A (November 2011). "Models for Golgi traffic: a critical assessment". Cold Spring Harb Perspect Biol. 3 (11): a005215. doi:10.1101/cshperspect.a005215. PMC 3220355. PMID 21875986.{{cite journal}}: CS1 maint: date and year (link), Drs. Glick and Luini analyze five different models for trafficking secretory cargo through the Golgi complex: (1) anterograde vesicular transport between stable compartments, (2) cisternal progression/maturation, (3) cisternal progression/maturation with heterotypic tubular transport, (4) rapid partitioning in a mixed Golgi, and (5) stable compartments as cisternal progenitors. Since the pathways and mechanisms of Golgi traffic remain heavily debated, Glick and Luini methodically describe each model in detail and list the strengths and weaknesses of each, acknowledging that no single model can easily explain all of the data on Golgi trafficking. With regard to selecting candidates for the “best” model, Glick and Luini approach this assessment from two different angles. First, from a mainly theoretical point of view, the authors try to assess which model might best reflect certain “core mechanisms” that are likely to be conserved in most or all eukaryotes; and based on this theoretical point of view, they favor Model 2 (cisternal progression/maturation). However, from a practical point of view, Glick and Luini focus their assessment on which model would serve as the best “working hypothesis” to guide future experimentation, and in this regard they cite Model 3 as the best candidate. In support of their assessment, Glick and Luini note that Model 2 is unable to explain all of the data, and at the same time there is “growing evidence” that heterotypic tubular connections are important for Golgi trafficking. Accordingly, Glick and Luini Glick BS, Luini A (November 2011). "Models for Golgi traffic: a critical assessment". Cold Spring Harb Perspect Biol. 3 (11): a005215. doi:10.1101/cshperspect.a005215. PMC 3220355. PMID 21875986.{{cite journal}}: CS1 maint: date and year (link)conclude that Model 2’s “core mechanism” theory can be “elaborated” by means of heterotypic tubular transport between cisternae”, and thus Glick and Luini select Model 3 as the best working hypothesis to guide future experimentation.

Wang et al., Biochem. 46(51):14751-14761(2007)

In a 2007 article,Wang S, Engohang-Ndong J, Smith I (December 2007). "Structure of the DNA-binding domain of the response regulator PhoP from Mycobacterium tuberculosis". Biochemistry. 46 (51): 14751–61. doi:10.1021/bi700970a. PMC 2535579. PMID 18052041.{{cite journal}}: CS1 maint: date and year (link) CS1 maint: multiple names: authors list (link) found that in its crystal structure, the PhoP protein exists as a hexamer, and Wang et al. speculated that the hexamer is probably not seen in cells because the hexamer structure blocks the recognition helix (thus preventing it from interacting with the major groove of DNA). In this regard, it should be noted that if the PhoP protein were a hexamer and could only bind the minor groove of DNA, specificity would lost in this minor groove DNA-binding mode. As shown in Figure 5 of Wang et al., Biochem. 46(51):14751-14761(2007), most of the positively charged surfaces (shown in blue in Figure 5) are exposed in the case of the hexamer structure (including the wing residues, which, according to the authorsWang S, Engohang-Ndong J, Smith I (December 2007). "Structure of the DNA-binding domain of the response regulator PhoP from Mycobacterium tuberculosis". Biochemistry. 46 (51): 14751–61. doi:10.1021/bi700970a. PMC 2535579. PMID 18052041.{{cite journal}}: CS1 maint: date and year (link) CS1 maint: multiple names: authors list (link), play an important role in DNA binding by interacting with the minor groove of DNA). In this way, multiple molecules of PhoPC are able to bind DNA through interactions with minor grooves, but specificity of DNA sequence-specific binding is lost.

Week 6 -- Selecting an Article Topic

Article Selection Rationale

Group 82H has decided to improve the article for XRCC4. As indicated in the current Wikipedia stub, as well as in our Watson text book, XRCC4 is a DNA repair protein encoded by the XRCC4 gene in humans; and in particular, the XRCC4 protein functions in conjunction with DNA ligase IV and the DNA-dependent protein kinase in the repair of DNA double-strand break by non-homologous end joining and the completion of V(D)J recombination events. In addition to the fact that XRCC4 is covered in our textbook, a review of the scientific literature reveals an abundance of very recent articles discussing the importance of XRCC4 and reporting on current research in this area. This is in fact, a very exciting time for XRCC4, as a relatively recent study by Hammel, et al., 2011, revealed the crystal structure of the XLF-XRCC4 complex. Specifically, a "key lock" interaction and hydrogen bonds that holds both proteins together to maintain the complex required for ligation of the DSB, characterization of the C-terminal domain shows the key structures responsible for DNA binding in a concentration-dependent way, and another DNA binding region at the XLF-XRCC4 interface. These findings are extremely critical in the development of new therapeutics as they can serve as target sites to ensure that the "proper alignment of damaged DNA for ligation by DNA Ligase IV is successfully completed. Other recent research topics also include examination of the potential linkage between XRCC4 and cancers such as bladder cancer and hepatitis B-associated hepatocellular carcinoma.

With regard to specific ideas for improving the XRCC4 Wikipedia article, the current stub has minimal text (less than 280 words), and despite the abundance of recent scientific publications that we found (many with publication dates within the past several months), all of the references listed in the current stub — except for two― are over a decade old; and the two “newer” articles are from 2007 and 2008. With regard to specific topics for us to research and include in the improved XRCC4 article, we note that current stub has no background section, no explanation as to how double-stranded breaks can occur, and limited explanation of the pathway for DNA repair for this type of damage. We could also cover topics such as the XRCC4 gene, XRCC4 protein structure, other proteins working in concert with XRCC4, schematics of the mechanism, and descriptions as to how mutations occurring in different splice variants of this protein (and DNA sequences associated with these mutations) are the basis of specific diseases. We could also cover experimental techniques used to determine mutations, protein homology, and we could include substantial coverage on the recent publications on XRCC4.

Week 7 Wiki Assignment for Team 82H (XRCC4) -- Starting Work on the Article

List of Potential References for the XRCC4 Article

• Wu CN, Liang SY, Tsai CW, Bau DT, The role of XRCC4 in carcinogenesis and anticancer drug discovery, Recent Pat Anticancer Drug Discov. 2008 Nov;3(3):209-19. Review. PMID 18991789.

•• Important Review for role of XRCC4 in cancer

• Dahm K, Role and regulation of human XRCC4-like factor/cernunnos, J Cell Biochem. 2008 Aug 1;104(5):1534-40. doi: 10.1002/jcb.21726. Review. PMID 18335491

•• Important Review for role of XLF

• Li Z, Alt FW, Identification of the XRCC4 gene: complementation of the DSBR and V(D)J recombination defects of XR-1 cells, Curr Top Microbiol Immunol. 1996;217:143-50. Review. PMID 8787623

•• Important Review for the XRCC4 gene

• Andres et al., A human XRCC4-XLF complex bridges DNA, Nucleic Acids Res. 2012 Feb; 40(4):1868-78. Epub 2012 Jan 27. PMID 22287571 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3287209/

•• Evidence is presented for how XRCC4-XLF complexes robustly bridge DNA molecules: DNA Ligase IV-independent bridging activity by XRCC4-XLF suggests an early role for this complex during end joining.

• Wu et al., Non-homologous end-joining partners in a helical dance: structural studies of XLF-XRCC4 interactions, Biochem Soc Trans. 2011 Oct;39(5):1387-92, suppl 2 p following 1392. doi: 10.1042/BST0391387 PMID 21936820.

•• Individual crystal structures show that the dimeric proteins XRCC4 and XLF are homologues with protomers containing head domains and helical coiled-coil tails related by approximate two-fold symmetry

•• Biochemical, mutagenesis, biophysical and structural studies have identified the regions of interaction between the two proteins and suggested models for the XLF-XRCC4 complex.

• Cottarel, J; et al. (2013). "A non-catalytic function of the ligation complex during non homologous end joining". Journal of Cell Biology 200 (2): 173 - 186. http://www.ncbi.nlm.nih.gov/pubmed/23345432

• Hammel, Michal, et al. (2011). "Crystal Structure of XLF-XRCC4 Complex Provides Model for Double-Strand Break Repair". Journal of Biological Chemistry 286: 32638 - 32650. http://www.jbc.org/content/286/37/e99966.full

• Watson, James D., et al. (2008). Molecular Biology of the Gene. 6th Edition. New York: Cold Spring Harbor Laboratory Press. pp. 275 - 278.

• Yurchenko, Vyacheslav, et al. (2006). "SUMO Modification of Human XRCC4 Regulates Its Localization and Function in DNA Double-Strand Break Repair". Molecular Cell Biology 26(5): 1786 - 1794. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1430232/

• Jung et al., Polymorphisms of DNA repair genes in Korean hepatocellular carcinoma patients with chronic hepatitis B: possible implications on survival, J Hepatol. 2012 Sep;57(3):621-7. doi: 10.1016/j. jhep.2012.04.039. Epub 2012 May 29, PMID 22659345

•• Conclusion: Polymorphisms of DNA repair genes play a potential role in the development, progression, and survival of Korean HCC patients with chronic HBV infection.

• Mahaney et al., XRCC4 and XLF form long helical protein filaments suitable for DNA end protection and alignment to facilitate DNA double strand break repair, Biochem Cell Biol. 2013 Feb;91(1):31-41. doi: 10.1139/bcb-2012-0058. Epub 2013 Feb 5. PMID 23442139.

• Zhang et al, Effects of expression level of DNA repair-related genes involved in the NHEJ pathway on radiation-induced cognitive impairment, J Radiat Res. 2013 Mar 1;54(2):235-42. doi: 10.1093/jrr/rrs095. Epub 2012 Nov 7. PMID 23135157

• Zhou LP, Luan H, Dong XH, Jin GJ, Ma DL, Shang H, Association of functional polymorphisms of the XRCC4 gene with the risk of breast cancer: a meta-analysis, Asian Pac J Cancer Prev. 2012;13(7):3431-6. PMID 22994773

• Zheng Z, Ng WL, Zhang X, Olson JJ, Hao C, Curran WJ, Wang Y., RNAi-mediated targeting of noncoding and coding sequences in DNA repair gene messages efficiently radiosensitizes human tumor cells, Cancer Res. 2012 Mar 1;72(5):1221-8. doi: 10.1158/0008-5472.CAN-11-2785. Epub 2012 Jan 11. PMID 22237628.

• Mandal RK, Singh V, Kapoor R, Mittal RD, Do polymorphisms in XRCC4 influence prostate cancer susceptibility in North Indian population?, Biomarkers. 2011 May;16(3):236-42. doi: 10.3109/1354750X.2010.547599. PMID 21506695.

•• XRCC4 and bladder cancer.

• Mittal RD, Gangwar R, Mandal RK, Srivastava P, Ahirwar DK, Gene variants of XRCC4 and XRCC3 and their association with risk for urothelial bladder cancer, Mol Biol Rep. 2012 Feb;39(2):1667-75. doi: 10.1007/s11033-011-0906-z. Epub 2011 May 27. PMID 21617942

•• linking XRCC4 to bladder cancer. Jgould1400 (talk) 06:16, 12 March 2013 (UTC)

• Q13426 (XRCC4_Human). UniProt. Last modified 3/6/13.

-Contains gene and protein names, protein composition and subunits (motif domains), amino acid modifications, isoforms. http://www.uniprot.org/uniprot/Q13426

• Junop, Murray S., et al., (2000), “Crystal structure of the Xrcc4 DNA repair protein and implications for end joining”, The EMBO Journal 19: 5962 – 5970. PMID 11080143.

http://www.nature.com/emboj/journal/v19/n22/abs/7593410a.html -Findings of crystal structure of functional fragment of protein composing of a dumb-bell-like tetramer, N-terminal domain are globular and is a potential DNA binding domain, its C-terminal stalk may serve a role in interacting with ligase IV; all required for effective DNA repair in NHEJ.

• Hammel, M., et al., (2011), “XRCC4 protein interactions with XRCC4-like factor (XLF) create an extended grooved scaffold for DNA ligation and double strand break repair”, J Biol Chem 286(37): 32638-50. PMID 21775435. http://www.ncbi.nlm.nih.gov/pubmed/21775435

• Li, Y., et al., (2008), “Crystal structure of human XLF/Cernunnos reveals unexpected differences from XRCC4 with implications for NHEJ”, EMBO J 27(1):290-300. PMID 18046455.

http://www.ncbi.nlm.nih.gov/pubmed/18046455 -Authors use various biochemical and biophysical techniques to elucidate the functional motif of Cer-XLF as a short, four helical bundle and a C-terminal helical structure inserted between its coiled-coil and head domain. They also predict the stoichiometric ratios of the XRCC4:XLF:Ligase IV complex.

• Wu, Pei-Yu, et al., (2009), “Structural and Functional Interaction between the Human DNA Repair Proteins DNA Ligase IV and XRCC4”, Molecular and Cellular Biology 29(11):3163. PMID 19332554.

http://mcb.asm.org/content/29/11/3163.full.pdf+html -Authors characterized specific protein domains and the structural and functional interactions between XRCC4, LigIV and Cernunnos XRCC4-like Factor (Cer-XLF) in the complex required for ligation of DSB during NHEJ. Found the central domain of XRCC4 contains a helix-loop-helix motif that is the binding interface for the inter-BRCT linker at the C-terminus of LigIV.

• Mari, PO, et al., (2006), “Dynamic assembly of end-joining complexes requires interaction between Ku70/80 and XRCC4”, Proc Natl Acad Sci USA 103(49):18597-602. PMID 17124166.

http://www.pnas.org/content/103/49/18597.full.pdf+html -Generation of DSB using near IR laser pulses demonstrate accumulation of NHEJ proteins, XRCC4/ligase IV in the presence of Ku70/80 in irradiated regions in vivo. Authors elucidate the function and assembly of the complex involving these proteins. Cdunca12 06:29, 15 March 2013 (UTC)

Draft Outline for XRCC4 Article

1. Lead

-Thorough but concise overview of XRCC4 protein: its role in the non-homologous end joining (NHEJ) pathway; importance of repairing double-stranded DNA breaks (DSB) prior to replication; basic mechanism with Ku70/80, DNA-PKcs and Artemis, complex of XRCC4-Cer-XLF-LigIV proteins; diseases associated with mutations in XRCC4; recent or current debates or controversies in research or clinical fields on the topic.

2. Structure

- Include a "Gene and Protein" section with fundamental information such as the chromosomal location of the gene, exons, protein splice variants and structure and domains (include appropriate images)

3. Function

-Review of the various types of DNA repair mechanisms and the type of DNA damage each corrects. Description of the two types of DSB: post-replication (using sister chromosome as the template to fill in the DSB) and pre-replication (NHEJ).

4. Mechanism / Biological Processes / Location within the cell

- Include in-depth discussion of the specific mechanism of the protein and other proteins involved in the pathway with a clear schematic. Include description of structure of the binding sites of XRCC4 and associated NHEJ proteins and how they specifically interact with DNA and with each other.

- The localization of XRCC4 and associated proteins in the nucleus and their gene expression and protein regulation as required for DSB repair.

-Compare mechanism between eukaryotes and prokaryotes.

5. Pathology

- Known mutations and diseases they cause

- Possibilities for diagnosis

- Possibilities for prevention

- Possibilities for treatment

6. Interactions

7. Experimental techniques for studying the gene and protein

-A few key methods described in the articles referenced and how the authors determined their conclusions about protein structure and regulation.

-Most current methods used to determine DNA sequence to uncover mutations or hot spots belonging to specific individuals for development of personalized medicine.

8. History section

- Who discovered the protein and how?

- Genetics

9. References

10. Further reading

11. External links

Cdunca12 19:41, 17 March 2013 (UTC)

Potential Images

PDB 1xnt EBI

File:Mechanism of non-homologous end joining for double-stranded DNA breaks MF.png

Mechanism of non-homologous end joining for double-stranded DNA breaks MF.png

Cdunca12 04:43, 21 March 2013 (UTC)

Progress Report for Units 11-12

A very substantial amount of new material was added since our last progress report, including the new section on "History and Identification of the XRCC4 Gene", the new section on "Anti-XRCC4 Antibodies", and the new subsection on "Endometriosis Susceptibility" that was added to the Pathology section; and in addition, information and content was consolidated in several locations and many references were added. We have also considered the various comments we received and have made revisions and additions where appropriate. Jgould1400 (talk) 04:36, 26 April 2013 (UTC)

Progress Report for Units 13-14 (Final Project Report)

Substantial revision/supplementation of several sections/subsections; addition of some subsections and an image; addition of text and several references; searched for more information and to see if we’ve missed anything significant; and corresponded with reviewers. OA Keilana said that the article looks great. Jgould1400 (talk) 21:37, 11 May 2013 (UTC)