Speaker: Dr. José Alcamí
Instituto de Salud Carlos III – Madrid, Spain

Organizer: IRB Barcelona 
Host: Antonio Celada, IRB Barcelona
Date: Friday, 27 January 2012, 12:00h.
Place: Aula Fèlix Serratosa, Parc Cièntific de Barcelona, Spain

Abstract:

Background
Tat protein of HIV-1 is essential for efficient elongation of viral transcripts and contributes to viral transcription in concert with other regulatory elements binding the HIV-LTR. Tat encoding gene consists of two spliced exons separated in the HIV-1 genome by more than 2.300 nucleotides. First 72 residues at the N-terminal encoded by the first exon appear to be sufficient for viral RNA elongation and transcriptional activity, but the integrity of the coding second exon in most lentiviral genomes suggests that it should have some biological importance. In this work, we analyze the potential role of the second exon of Tat in the regulation of gene expression and functional biology of CD4+ T lymphocytes.
Materials and methods
Jurkat cells with inducible expression of intracellular full-length Tat (1-101aa) or a truncated form lacking the second exon (1-72aa) of Tat. Whole human genome and microRNA microarrays were used to analyze the role of different versions of Tat in gene expression and deregulation. According to these results the following cellular functions were analyzed: a) expression of membrane receptors linked to lymphocyte activation b) cellular polarization and chemotaxis c) Apoptosis induced by CD95 ligands and transanduction pathways involved.
Results
More than 1.000 genes were deregulated in Jurkat Tat101, whereas less than 300 genes were deregulated in Jurkat Tat72 (q-value <5%; fold change >2 or <-2). Ontological analysis indicated that several functions were impaired mainly in Jurkat Tat101 as cellular movement, growth and proliferation, cell-to-cell signaling, molecular transport, cell death, morphology, and T-cell activation. In accordance with changes in gene expression Tat101 induced changes in cell polarization, cytoskeletal structure and chemotaxis capacity. Besides, a strong resistance to apoptosis induced through CD95 was observed in Tat101 cells. Accordingly to apoptosis resistance, activity of caspases 3, 8 and 9 was reduced in Tat101-expressing Jurkat cells whereas Bcl2 levels and c-FLIP-s isoform were increased. These alterations were attenuated or absent in Jurkat Tat72 cells.
Conclusion
Expression of full-length Tat protein produced dramatic structural changes and impaired essential functions in CD4+T cells as compared to Tat72 (first exon) expression. Consequently, although first exon is transcriptionally autonomous, second exon appears to be necessary for triggering HIV-1 pathogenic events induced by Tat.