B lymphocytes respond to antigen by proliferating and differentiating into antibody-secreting plasmacytes. That response can be enhanced or inhibited by antigen-specific T-helper (TH) or T-suppressor (TS) cells, respectively. The TNP-specific IgA?315?-secreting MOPC-315 myeloma plasma cells arise by differentiation from malignant small, nonsecretory lymphocytoid stem cells. The proliferation and differentiation of those stem cells can be regulated by distinct subsets of carrier-specific TH and TS cells. I was interested in determining the biochemical changes involved in activation and suppression of MOPC-315 cell secretory differentiation. Until purified helper and suppressor regulatory factors were available from my monoclonal differentiation helper and suppressor T-cell lines, I investigated; (1) whether MOPC-315 differentiation could be modulated by the thymus-independent antigen TNP-LPS (E. coli 055:B5 lipopolysaccharide), and (2) what some of the metabolic requirements were for that modulation. I found that while unconjugated or FITC-conjugated LPS had no effect on 315 cell differentiation when added at doses lower than 10 micrograms/ml in vitro, significant enhancement of 315 cell PFC frequency was observed with from 0.001-0.1 micrograms/ml TNP-LPS and significant inhibition of 315 cell anti-TNP IgA PFC frequency was induced at 1.0 micrograms/ml TNP-LPS. These low dose effects were blocked if monoclonal anti-idiotype?315? antibody or DNP-glycine were included in the culture media implying that TNP-LPS engagement of surface membrane IgA?315? was required. The stimulatory dose of TNP-LPS had to be present for only the first 2 hrs to get maximal enhancement measured 48 hrs after culture initiation. The inhibitory dose of TNP-LPS had to be present 24 hrs to get maximal inhibition of PFC frequency and maximal unresponsiveness to helper T-cell stimulation of secretory differentiation. This unresponsiveness once induced lasted 5-7 days. Within the first 1 hr of culture with the stimulatory dose of TNP-LPS, a serine esterase is activated which is required for the PFC frequency enhancement. This serine esterase activity requires phospholipid methylation within the first 15 min of TNP-LPS incubation to be induced. The inhibitory dose of TNP-LPS does not induce these changes but alters the 315 cell metabolism such that subsequent culture with 0.01 micrograms/ml TNP-LPS does not enhance PFC frequency and the serine esterase and phospholipid methylation does not occur. I am continuing these studies and am beginning to determine temporal changes in nuclear, cytoplasmic, and membrane protein phosphorylation using 2D gel electrophoresis. (LB)

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA028708-06
Application #
3168294
Study Section
Allergy and Immunology Study Section (ALY)
Project Start
1980-07-01
Project End
1987-02-28
Budget Start
1986-03-01
Budget End
1987-02-28
Support Year
6
Fiscal Year
1986
Total Cost
Indirect Cost
Name
University of South Alabama
Department
Type
Schools of Medicine
DUNS #
City
Mobile
State
AL
Country
United States
Zip Code
36688