Title

Differential Regulation of Soluble and Membrane CD40L Proteins in T Cells: Implication of ADAM-10

Publication Date

4-2006

Type of Culminating Activity

Thesis

Degree Title

Master of Science in Biology

Department

Biology

Major Advisor

Denise Wingett

Advisor

Cheryl Jorcyk

Advisor

Julia Oxford

Abstract

CD40 ligand (CD40L) is an important immunoregulatory protein expressed by activated T cells. This protein exists as two isoforms, a 33 kDa membrane glycoprotein and a truncated 15-18 kDa soluble form. In this report, it is demonstrated that soluble CD40L (sCD40L) and membrane CD40L are differentially regulated depending upon the activation stimulus. In T cell receptor (TCR)-activated cells, both soluble CD40L and membrane CD40L are expressed, and CD28 costimulation further increases their expression. The dissection of TCR generated signals into calcium and PKC-dependent pathways demonstrates that calcium ionophore is sufficient to induce membrane CD40L yet insufficient for sCD40L production. In contrast, PKC-dependent signals result in a preferential release of sCD40L. Moreover, sCD40L production is blocked by the ZN+2-dependent matrix metalloprotease inhibitors, TAPI-2 and CM 6001, while membrane CD40L expression is concurrently increased. The profile of a PKC-dependent proteolytic event involving a metalloprotease suggested the potential involvement of ADAM-10 or TNF-α converting enzyme (TACE) proteases in the generation of sCD40L. Western blot experiments demonstrate that recombinant ADAM-10 cleaves CD40L protein to generate an 18 kDa form corresponding to sCD40L while TACE-mediated cleavage of CD40L was not observed. These results indicate that sCD40L production is an active process regulated by a PKC-dependent pathway that involves the proteolytic cleavage by ADAM-10 or a related metalloprotease. Given the role of sCD40L in numerous disease pathologies and its cytokine-like ability to activate both proximal and distal immune responses, the regulated cleavage of this protein can be expected to affect disease progression or outcome.

Files over 30MB may be slow to open. For best results, right-click and select "save as..."

Share

COinS