Evidence of Rab3A expression, regulation of vesicle trafficking, and cellular secretion in response to heregulin in mammary epithelial cells

Heregulin beta1 (HRG), a combinatorial ligand for human growth factor recep tors 3 and 4, is a regulatory polypeptide that promotes the differentiation of mammary epithelial cells into secretory lobuloalveoli. Emerging evidenc e suggests that the processes of secretory pathways, such as biogenesis and trafficking of vesicles in neurons and adipose cells, are regulated by the Rab family of low-molecular-weight GTPases. In this study, we identified R ab3A as a gene product induced by HRG. Full-length Rab3A was cloned from a mammary gland cDNA library. We demonstrated that HRG stimulation of human b reast cancer cells and normal breast epithelial cells induces the expressio n of Rab3A protein and mRNA. in a cycloheximide-independent manner. HRG-med iated induction of Rab3A expression was blocked by an inhibitor of phosphat idylinositol 3-kinase but not by inhibitors of mitogen-activated protein ki nases p38(MAPK) and p42/44(MAPK). Human breast epithelial cells also expres s other components of regulated vesicular traffic, such as rabphilin 3A, Do c2, and syntaxin. Rab3A was predominantly localized in the cytosol, and HRG stimulation of the epithelial cells also raised the level of membrane-boun d Rab3A. HRG treatment induced a profound alteration in the cell morphology in which cells displayed neuron-like membrane extensions that contained Ra b3A-coated, vesicle-like structures. In addition, HRG also promoted the sec retion of cellular proteins from the mammary epithelial cells. The ability of HRG to modify exocytosis was verified by using a growth hormone transien t-transfection system. Analysis of mouse mammary gland development revealed the expression of Rab3A in mammary epithelial cells. Furthermore, expressi on of the HRG transgene in Harderian tumors in mice also enhanced the expre ssion of Rab3A. These observations provide new evidence of the existence of a Rab3A pathway in mammary epithelial cells and suggest that it may play a role in vesicle trafficking and secretion of proteins from epithelial cell s in response to stimulation by the HRG expressed within the mammary mesenc hyma.