The way in which a cell regulates the synthesis and assembly of its membrane is fundamental question in cell biology. It is attractive to speculate that cells have evolved mechanisms designed to coordinate the synthesis of membrane proteins. These mechanisms would operate to maintain homeostasis during normal cell growth and induce the synthesis of a required cohort of membrane proteins during periods of cell differentiation. The general goal of my research is to identify and elucidate these regulatory pathways. This proposal focuses specifically on the regulation of the synthesis and assembly of the endoplasmic reticulum (ER) during bacterial lipopolysaccharide (LPS)-induced differentiation of B- lymphocytes. Recent results from several laboratories, including our own, have shown that members of two different """"""""stress"""""""" protein families, GRP78/BiP and GRP94/ERp99, are abundant components of the ER. In addition, it has recently been demonstrated that GRP78/BiP binds to incorrectly or incompletely folded newly made proteins and that this binding stimulates the transcription of BiP mRNA. Based upon these findings, we have developed the working hypothesis that one of the normal roles of the stress proteins in the lymphocyte ER is to function as part of a system which monitors the level of immunoglobulin (Ig) in the ER lumen. As Ig synthesis increases, these proteins would then serve as a part of the regulatory pathway that tells the cell that the secretory load is increasing at that it must make more ER. The first specific aim of this proposal describes experiments designed to obtain the basic information necessary to establish the kinetic parameter of the ER differentiation process and to test key predictions of our model. The second specific aim is focused on developing an experimental system which will be used to provide a rigorous test of the model and to investigate specific molecular events predicted by the model. The results obtained from these studies should provide new useful insights into an important biological question. The findings would be of specific interest to the field of B-lymphocyte biology and of general interest to the field of membrane biogenesis.
