To identify potential SOX10 target genes, we will analyze the effects of altered SOX10 expression on the expression of multiple genes simultaneously using cDNA expression microarrays. Successful microarray experiments require: 1) appropriate mRNA populations to compare, 2) appropriate cDNA populations on the array, 3) critical secondary screens, and 4) defined functional assays. 1) We will compare expression patterns from cell cultures with altered SOX10 function. These will include primary cultures of SOX10Dom/+ versus +/+ neural crest and NC-Ms as well as cell lines transduced with SOX10 expressing viruses. 2)We are developing a standard set of mouse cDNA clones to use in microarray expression analyses based on the WASHU-Merck mouse ESTs. However, we have found that these generalized arrays may not be the most efficient source of cDNA for a particular tissue source. Therefore, we developed a novel strategy to isolate a set of cDNA clones representing the majority of genes that regulate NC-M development and function (Loftus et al, 1999). We will use this set for expression profiling in collaboration with Dr. Trent and the NHGRI microarray core. 3) Since we are screening for target genes of SOX10 transcriptional regulation, we will assess candidates by in situ hybridization using mutant SOX10 embryos at the time when SOX10 expressing cells are present but failing to migrate. Genes exhibiting a NC expression pattern in WT but not in mutant SOX10 expressing cells will be analyzed further. 4) Most in vivo approaches for defining gene function (transgenic and knockout strategies) require extensive cloning, viable offspring, and animal breeding. We will use our NC infection strategy described above (RCAS) to misexpress candidate gene products in NC and examine the effects on development of specific lineages. We have already identified one gene that results in a 4-fold increase in melanocyte number when melanoblasts are specifically targeted. - biotechnology research, cancer research, digestive diseases, gene mapping(non-human), neuroscience, pediatric research

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Intramural Research (Z01)
Project #
1Z01HG000136-02
Application #
6433677
Study Section
(GDRB)
Project Start
Project End
Budget Start
Budget End
Support Year
2
Fiscal Year
2000
Total Cost
Indirect Cost
Name
Human Genome Research
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Watkins-Chow, Dawn E; Varshney, Gaurav K; Garrett, Lisa J et al. (2017) Highly Efficient Cpf1-Mediated Gene Targeting in Mice Following High Concentration Pronuclear Injection. G3 (Bethesda) 7:719-722
Silver, Debra L; Hou, Ling; Somerville, Robert et al. (2008) The secreted metalloprotease ADAMTS20 is required for melanoblast survival. PLoS Genet 4:e1000003
Buac, Kristina; Pavan, William J (2007) Stem cells of the melanocyte lineage. Cancer Biomark 3:203-9
Matera, Ivana; Cockroft, Jody L; Moran, Jennifer L et al. (2007) A mouse model of Waardenburg syndrome type IV resulting from an ENU-induced mutation in endothelin 3. Pigment Cell Res 20:210-5
Baxter, Laura L; Hsu, Benjamin J; Umayam, Lowell et al. (2007) Informatic and genomic analysis of melanocyte cDNA libraries as a resource for the study of melanocyte development and function. Pigment Cell Res 20:201-9
Brooks, Brian P; Larson, Denise M; Chan, Chi-Chao et al. (2007) Analysis of ocular hypopigmentation in Rab38cht/cht mice. Invest Ophthalmol Vis Sci 48:3905-13
Riazuddin, Saima; Khan, Shaheen N; Ahmed, Zubair M et al. (2006) Mutations in TRIOBP, which encodes a putative cytoskeletal-organizing protein, are associated with nonsyndromic recessive deafness. Am J Hum Genet 78:137-43
Hakami, Ramin Mollaaghababa; Hou, Ling; Baxter, Laura L et al. (2006) Genetic evidence does not support direct regulation of EDNRB by SOX10 in migratory neural crest and the melanocyte lineage. Mech Dev 123:124-34
Hou, Ling; Arnheiter, Heinz; Pavan, William J (2006) Interspecies difference in the regulation of melanocyte development by SOX10 and MITF. Proc Natl Acad Sci U S A 103:9081-5
Silver, Debra L; Hou, Ling; Pavan, William J (2006) The genetic regulation of pigment cell development. Adv Exp Med Biol 589:155-69

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