Release, uptake, and effects of extracellular human immunodeficiency virus type 1 Tat protein on cell growth and viral transactivation.

We investigated whether during acute infection of T cells by human immunodeficiency virus type 1, tat encodes an early protein is released at concentrations sufficient to stimulate HIV-1 gene expression in a paracrine fashion and the events associated with the release of biologically active protein by cells transfected with the tat gene

B Ensoli

2020

Scholarcy highlights

  • During acute human immunodeficiency virus type 1 infection or after transfection of the tat gene, tat encodes an early protein protein is released into the cell culture supernatant
  • We investigated whether during acute infection of T cells by HIV-1, Tat is released at concentrations sufficient to stimulate HIV-1 gene expression in a paracrine fashion and the events associated with the release of biologically active protein by cells transfected with the tat gene
  • In order to determine whether during acute HIV-1 infection released Tat was able to transactivate HIV-1 gene expression in a paracrine fashion, Conditioned media from H9-infected T cells or from uninfected cells from sequential time points postinfection were assayed for transactivating activity by scrape loading into COS-1 cells transfected with the HIV-1 long terminal repeat-chloramphenicol acetyltransferase construct
  • To verify whether the activation of HIV gene expression by CM was due to the presence of Tat in the CM, H9-HIV CM was scrape loaded into cells transfected with a deletion mutant of the Tatresponsive region of the HIV-1 LTR
  • We have shown that during acute HIV-1 infection or after transfection of the gene, Tat is released into the cell culture medium and can stimulate maximal AIDS-Kaposi's sarcoma cell growth and low to moderate levels of HIV-1 gene expression
  • After being electroshocked, cells were incubated on ice for 8 to 10 min and transferred very gently to a 75-cm3 flask containing growth medium at 37°C. By this technique, more than 99% of the cells were alive and attached to the flasks 10 to 14 h after transfection
  • The transfection method utilized in our studies is characterized by high expression of Tat and may mimic this early phase of HIV-1 replication, when high Tat expression results in accumulation in the cytoplasm and release into the medium
  • Viral transactivation by extracellular tat encodes an early protein can occur in a situation of close cell proximity or contact , these results suggest that the cell growth-promoting activity may be more biologically relevant than the virus-transactivating effect of extracellular Tat. The experiments with purified Tat protein show that maximal cell growth is obtained with Tat concentrations much lower than those required to induce HIV transactiva-

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