The long-term goals of this project are to identify external factors and to isolate regulatory gene elements which control 1). the initiation of IgD expression during B cell development and 2). high rate IgM secretion after B cell activation. Since the gene for the constant region of the delta heavy chain is located downstream of Cmu, transcription of Cdelta is regulated by the extent of RNA polymerase readthrough beyond the termination site of the mu gene. Similarly, the transition of B lymphocytes to a state of high rate production of mRNA for secreted IgM is regulated by alteration of the termination site of the mu transcription unit to more upstream positions which precludes the synthesis of mRNA for mu chain of membrane IgM. In order to understand the mechanisms underlying these alterations in RNA polymerase termination, gene sequences which regulate the termination will be identified by transient transfection of constructs containing the putative termination sequences. The role of RNA polymerase will be investigated by substituting promoter elements such that the gene can be transcribed by RNA polymerase III. Efforts will be also made to analyze the regulation of termination in an in vitro RNA transcription system using crude extracts from different cell sources. In addition, the signals which induce early B lymphocytes to begin IgD expression will be identified in order to correlate external stimuli with the intracellular events. Finally, using the well characterized, factor responsive lymphoma line, BCLl, the genes which may be involved in induction of termination of RNA polymerases in yet more upstream locations late in B cell differentiation will be isolated by means of subtractive hybridization. These experiments, designed to dissect the elements which modulate RNA polymerase termination, should lead to better understanding of the regulation of Ig synthesis in particular, and further insights into this mode of gene regulation in general.
| Yuan, D; Goldings, E A (1988) Transient expression of pentameric IgM on the surface of NZB B cells. Mol Immunol 25:323-8 |
