Identification and functional evaluation of proteins controlling proliferation and differentiation of mammary epithelial cells
Final Report Abstract
The mammary gland represents a unique model-system to study gene function in adult stem cells. Its small fraction of stem cells (MSCs) can regenerate a functional mammary gland upon cleared fat pad transplantation. We optimized lentiviral transduction of mammary epithelial cells (MECs) to genetically modify non-enriched MSCs ex vivo. Transplantation of mixed MECs expressing different fluorescent proteins resulted in separate unicolored ducts probably originating from individual MSCs. We also genetically modified MSCs to gauge Stat5 function during mammary gland regeneration and differentiation. Stat5-knockdown did not affect primary ductal outgrowth but impaired side-branching and the emergence of mature alveolar cells from luminal progenitors. Conversely, the expression of a constitutively active variant of Stat5 (cS5-F) caused epithelial hyperproliferation, thickening of the ducts and precocious development of alveoli in virgin mice. Persistent expression of cS5-F during involution blocked apoptosis and caused formation of adenocarcinomas with short latencies. Microarray analysis revealed known and novel Stat5 target genes in non-tumorigenic cS5-F glands and tumors. Keywords: mammary stem cells, lentiviral gene transduction, organ reconstitution, gene function analysis, Stat5 target genes, mammary tumor formation
