The sheep erythrocyte receptor (aka p50, T11 or CD2) is a T lymphocyte specific cell surface glycoprotein which mediates antigen, lectin and accessory cell independent T lymphocyte activation. Anti-CD2 antibodies have been shown to either induce, or block T cell activation in an IL-2 dependent manner. The goal of the sponsors laboratory is to clone CD2 in order to study the structure-function relationship of this important molecule. To that end, we have isolated 5 recombinant cDNA inserts from a lambda GT-11 library which (a) range from 1000- 1500 bp, (b) are cross-reactive by Southern analysis, (c) code for protein determinants reactive with a polyvalent anti-CD2 antisera and (d) hybridize to 1.7 and 1.3 kb RNA transcripts found only in CD2 positive T cells but not CD2 negative B or T cells. This award will be applied to verify the identity of our cDNA isolates as CD2 by detecting CD2 epitopes in COS-7 fibroblasts transfected with pcEXV-3-CD2 constructs. This vector contains the SV40 early promotor and enhancer sequences. We than plan on cotransfecting CD2 positive and negative leukemic lymphoid cell lines with pSV-2-neo and pcEXV-3-CD2 constructs.
Our aims are (a) the isolation of stable transfectants which express cell surface CD2, (b) demonstration by FACS analysis of intact CD2 epitopes in transfected cells and (c) assay of transfectants for CD2 dependent responsiveness to anti-CD2 antibodies, lectins and mitogens. If we are successful in isolating transfectants with functional CD2, we propose future experiments to (a) map functionally important CD2 epitopes such as T112 and T113, (b) determine by selective mutagenesis which structures are involved in CD2 mediated signal transduction, (c) if specific transmembrane mutations can alter the function of CD2 as seen for p185 (neu protein) and (d) whether transfectants exhibit IL-2 dependent CD2 mediated responsiveness.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08CA001427-03
Application #
3079913
Study Section
Special Emphasis Panel (SRC)
Project Start
1988-07-01
Project End
1991-06-30
Budget Start
1990-07-01
Budget End
1991-06-30
Support Year
3
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Tulane University
Department
Type
Schools of Medicine
DUNS #
City
New Orleans
State
LA
Country
United States
Zip Code
70118
Malter, J S; Hong, Y (1991) A redox switch and phosphorylation are involved in the post-translational up-regulation of the adenosine-uridine binding factor by phorbol ester and ionophore. J Biol Chem 266:3167-71
Sullivan, D; Malter, J S (1991) A rapid and inexpensive method for the addition of poly A coding tracts to transcription vectors. Biotechniques 11:614, 616
Carter, B Z; Malter, J S (1991) Regulation of mRNA stability and its relevance to disease. Lab Invest 65:610-21
Makni, H; Malter, J S; Reed, J C et al. (1991) Reconstitution of an active surface CD2 by DNA transfer in CD2-CD3+ Jurkat cells facilitates CD3-T cell receptor-mediated IL-2 production. J Immunol 146:2522-9
Lampertico, P; Malter, J S; Gerber, M A (1991) Development and application of an in vitro model for screening anti-hepatitis B virus therapeutics. Hepatology 13:422-6
Malter, J S; Gerber, M A (1991) The polymerase chain reaction for hepatitis B virus DNA. Hepatology 13:188-90
Thompson, H W; Malter, J S; Steinemann, T L et al. (1991) Flow cytometry measurements of the DNA content of corneal epithelial cells during wound healing. Invest Ophthalmol Vis Sci 32:433-6
Rondon, I J; MacMillan, L A; Beckman, B S et al. (1991) Hypoxia up-regulates the activity of a novel erythropoietin mRNA binding protein. J Biol Chem 266:16594-8
Gillis, P; Malter, J S (1991) The adenosine-uridine binding factor recognizes the AU-rich elements of cytokine, lymphokine, and oncogene mRNAs. J Biol Chem 266:3172-7
Lauer, S A; Malter, J S; Meier, J R (1990) Human papillomavirus type 18 in conjunctival intraepithelial neoplasia. Am J Ophthalmol 110:23-7

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