Ronald Jemmerson, PhD

Professor Emeritus

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Professor Emeritus

Research

Antigen-Antibody Interactions; Apoptosis

My laboratory contributed to the seminal study showing that cytochrome c (Cyt c) translocates from mitochondria to the cytoplasm during apoptosis and plays a key role in the activation of programmed cell death (Cell 86:147-157, 1996). One monoclonal antibody specific for the native protein was used to remove Cyt c, thus showing that this protein and not a contaminant was responsible for the pro-apoptotic activity and another monoclonal antibody specific for the carboxyl terminal dodecapeptide was used to show Cyt c translocation to the cytoplasm by Western blotting. Later we found that Cyt c is released from apoptotic cells as an intact protein (Cell Death Differ. 9:538-548, 2002). Consequently, Cyt c has been explored as a potential clinical biomarker for aberrant apoptosis in vivo.

By distinguishing monoclonal antibodies that recognize native versus denatured Cyt c we also identified an antibody recognizing an altered conformation of Cyt c that has biological relevance (Biochemistry 36: 3599-3609, 1999). The altered form arises when the sixth ligand to the heme moiety is lost, endowing the protein with peroxidase activity and the ability to translocate to the nucleus without activating apoptosis (PNAS 119: e2213432119, 2022). This monoclonal antibody is, thus, a useful tool to examine a potential role of Cyt c in the nucleus, possibly affecting gene expression.

While developing an immunoassay to detect Cyt c in serum we discovered that leucine-rich alpha-2-glycoprotein-1 (LRG1) binds Cyt c and inhibits its detection in an antibody-based assay (Apoptosis 11:1121-1129, 2006). LRG1 has been implicated as a biomarker for a variety of cancers and microbial infections. However, its function was unknown. We have shown that LRG1 binds Cyt c in a manner similar to that of the protein Apaf-1 that initiates the intrinsic pathway of apoptosis. In vitro serum LRG1 acts as a survival protein extending the half -life of lymphocytes (Apoptosis 15:139-152, 2010). Recently, we demonstrated that, in MCF-7 breast cancer cells, cytoplasmic LRG1 competes with Apaf-1 for binding Cyt c in situ and, thus, inhibits the onset of apoptosis. We developed an enzyme-linked immunosorbent assay for LRG1 that may be useful clinically (J. Immunol. Methods 336:22-29, 2008).

Our apoptosis-related research evolved from studies of antibody recognition employing Cyt c as the model antigen. We made a number of observations that give insight into the antibody response with implications for vaccine development. We uncovered an artifact in immunoassays that misled researchers into believing that peptides could be broadly used as vaccines (PNAS 84:9180-9184, 1987). Even when a peptide can bind antibodies elicited against a native protein antigen this does not necessarily indicate that the peptide can activate B cells producing those antibodies as a certain affinity threshold must be met (Mol. Immunol. 26:301-307, 1989). We also showed that approximately 10% of memory B cells can be activated in an environment of T cell help in the absence of the cognate B cell antigen (Eur. J. Immunol. 21:951-958). This may explain, at least in part, the longevity of B cell memory after antigen has been depleted in vivo. We found that affinity maturation in the antibody response to a protein antigen follows the paradigm established for responses to smaller chemical compounds. Thus, an affinity increase of several hundred fold can result from DNA mutations at only a few key nucleotide positions in antibody genes (J. Immunol. 157:5329-5338, 1996). Much more about this phenomenon is explained in a follow-up review (Recent Res. Dev. Immunol. 4:13-25, 2002).

A surprising observation was a higher frequency of B lymphocytes in mice specific for mouse Cyt c than for foreign Cyts c (Eur. J. Immunol. 25:784-791, 1995). The autoreactive B cells generally expressed the same antibody heavy and light chain genes but were quite variable in CDR3 of the heavy chain. We speculate that mouse Cyt c may serve as a ligand for B cell positive selection. Supporting this idea, the antibody response to the closely related pigeon Cyt c actually initiates as a response more specific for mouse Cyt c utilizing those same variable region genes and eventually becomes more specific for pigeon Cyt c (PNAS 92:12379-12383, 1995).