User:Emmnanuel Quiroz

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Contact Info

Emmanuel Quiroz (an artistic interpretation)

• Emmanuel Quiroz
• Biological Engineering
• 416 Beacon St
• Address 2
• Boston, MA, 02115
• equiroz AT mit DOT edu

I work in the Wittrup Lab at MIT. I learned about OpenWetWare from Class, and I've joined because To access my classes material.

Chernobyl Fungi Proposal

Topic

Recent studies have discovered fungal species that survive and proliferate in environments of ionizing radiation such as the Chernobyl reactor. These fungi have very efficient DNA repair gene systems that may be homologous to the human pathway. These fungal systems need to be further studied through microarrays and more site-specific anaylsis to a better understanding of how Chernobyl fungi are different. Use of this information can potentially be helpful in cancer research and in developing new therapeutic approaches.

Research Goals

To explore the evolution of the DNA repair mechanism in Chernobyl fungi Cladosporium sphaerospermum to determine differences in gene regulation for potential new approaches in cancer treatment.

1. Find an efficient DNA repair gene expression in Chernobyl Fungi
2. Compare the fungi system to that of similar fungi

Research Plan

Perform microarray analysis on Chernobyl fungi( Cladosporium sphaerospermum and Penicillium hirsutum ) and compare their gene expressions of DNA repair systems to the well know genome of yeast (Saccharomyces cerevisiae)

1. For each species, we will analyze three samples:
   • No treatment: Our overall control
   • Radiation treatment: Simulates conditions found in Chernobyl region (γ-rays)
   • MMS: An alternative DNA-damaging agent, will allow us to determine if the entire DNA pathway has been affected or whether simply radiation resistance genes have been affected
2. Important considerations:
   • Can we compare three samples on one microarray? (potentially three fluorescent dyes?)
   • How will we compare two different array chips? (new method of log2 analysis required? See References #13 for more analysis ideas.)

BLAST two species' sequences against each other

1. Compare differences in the sequences to microarray analysis result
2. This will help confirm that variations in microarray fluorescence accurately correlated to differences in the regulatory pathway or genetic sequence of the two fungi
3. Taxonomy NCBI taxonomy database
   • Saccharomyces cerevisiae: Eukaryota; Fungi; Dikarya; Ascomycota; Saccharomycotina; Saccharomycetes; Saccharomycetales; Saccharomycetaceae; Saccharomyces
   • Cladosporium sphaerospermum: Eukaryota; Fungi; Dikarya; Ascomycota; Pezizomycotina; Dothideomycetes; Dothideomycetidae; Capnodiales; Davidiellaceae; mitosporic Davidiellaceae; Cladosporium
   • Penicillium hirsutum: Eukaryota; Fungi; Dikarya; Ascomycota; Pezizomycotina; Eurotiomycetes; Eurotiomycetidae; Eurotiales; Trichocomaceae; mitosporic Trichocomaceae; Penicillium

Conduct more site-specific analysis to more intimately determine changes in one particular gene (See possible gene candidates below)

1. Either use "pre-determined" gene involved in DNA repair that we have explored in a paper (See References) or a gene found via our microarray
   • Experiment Ideas:
   • Potentially create a knockout of the gene and see how repair/cell cycle is affected
   • Apply some sort of agent to up- or down-regulate the gene

Potential Gene Targets

1. repair (RAD50, RAD51)
2. recombination (HRP1)
3. chromosome stability (CHL1, CTF4)
4. endocytosis (VID21)
5. ubiquitin degradation (GRR1)
6. transcription (BUR2)

References

1. Ionizing radiation: how fungi cope, adapt, and exploit with the help of melanin ----- Fungi are seen to proliferate in environments of ionizing radiation such as the Chernobyl reactor and on the outer shell of space craft. These fungi show an increase in melanin expression which suggest that melanin can function in energy capture and prevent DNA damage. It was also seen through microarray analysis that exposure to radiation caused an up regulation in DNA repair genes. Interestingly, many of the radiation resistance genes share significant homology with human genes that might be exploited in the development of novel radiation-based cancer therapies. Link
2. Fungi and Ionizing Radiation from Radionuclides ----- Radionuclides, atoms with unstable nuclei due to excess energy that undergo radioactive decay, have provided interesting clues as to gene regulation since the Chernobyl power plant accident. Researchers have focused on various forms of fungi as a model of these effects because its large surface area allow for greater uptake capacity of nutrients and make it preferential to surround host plants. Through broad microarray analysis of various fungal stains, short term exposure of yeast to gamma and X radiation has been shown to elicit the up-regulation of genes involved in DNA cell repair, cell rescue defense, and cell fate and metabolism. Cladosporium cladosporioides and Penicillium roseopurpureum (saprotrophic micro-fungi) in particular were shown to over grow on carbon-based radioactive debris, and while radionuclides were incorporated into the cytoplasm, they were highly absorbed in ion-exchange sites (i.e. the cell membrane). Additionally, intense radiation of soil fungal communities led to a dominance of melanized fungal species, indicating a potential radiological protective mechanism in the pigment. Dighton, John; FEMS Microbiol Lett 2008 Vol. 281 Iss. 2 p. 109 - 20ppublish Link
3. Chernobyl effect: growth characteristics of soil fungi Cladosporium cladosporioides (Fresen) de Vries with and without positive radiotropism -- Zhdanova, N Fomina, M Redchitz, T Olsson, S Polish Journal of Ecology [Pol. J. Ecol.]. Vol. 49 2001 Vol. 49 Iss. 4 p. 309 - 318 Link
4. Fungi from Chernobyl: mycobiota of the inner regions of the containment structures of the damaged nuclear reactor Zhdanova, NN Zakharchenko, VA Vember, VV Nakonechnaya, LT Mycological Research [Mycol. Res.]. Vol. 104 2000 Vol. 104 Iss. 12 p. 1421 - 1426 Link
5. Normal expression of DNA repair proteins, hMre11, Rad50 and Rad51 but protracted formation of Rad50 containing foci in X-irradiated skin fibroblasts from radiosensitive cancer patients (RAD51 in cancer cells) C Djuzenova, B Mühl, R Schakowski, U Oppitz and M Flentje British Journal of Cancer (2004) 90, 2356–2363 Link
6. Link (XP variant HR pathway)
7. Link (recombination proteins in yeast - helpful detailed descriptions of Rad50 and many other proteins)
8. Link (genes required for ionizing radiation resistance in yeast)
9. Functional genomics of the yeast DNA-damage Gerard Cagney, David Alvaro, Robert JD Reid, Peter H Thorpe, Rodney Rothstein and Nevan J Krogan, Genome Biology 2006, 7:233 Link
10. http://www.nature.com/nmeth/journal/v2/n5/full/nmeth756.html (Methods of Microarray Comparative Analysis)
11. Interactions of Fungi and Radionuclides in Soil John Dighton, Tatyana Tugay, and Nelli Zhdanova Soil Biology ISSN 1613-3382 Volume 13 Book Microbiology of Extreme Soils Link

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