Biomedical subjects

S Berchtold

Publications and source records attributed to S Berchtold.

22 records · Page 2Linked to original sources

PubMed1994

A single stem-loop structure within the HTLV-1 Rex response element is sufficient to mediate Rex activity in vivo.

The human T-cell leukemia virus (HTLV-1) Rex protein is required for the cytoplasmic expression of the incompletely spliced transcripts that encode the viral structural proteins. This effect is mediated by a highly structured cis-acting RNA element of 254 nucleotides termed the Rex response element, or RexRE. Here we demonstrate that one of the four known RexRE stem-loop structures as well as a 43-nt segment derived from this element is sufficient to mediate Rex function in vivo. Upon duplication, this stem-loop is shown to function as efficiently as the full-length RexRE. In vitro RNA binding analyses with wildtype and mutagenized RNA show that this stem-loop contains a high affinity binding site for Rex that coincides with a predicted bulge structure in the central part of this stem-loop. These results indicate that a small region of the RexRE containing a high affinity binding site is sufficient to mediate Rex function and suggest that sequences outside of this binding site have no unique role in mediating Rex regulation.

Base Sequence↗

PubMed1994

Exchange of functional domains between Rev proteins of HIV-1 and SIVmac239 results in a dominant negative phenotype.

The Rev proteins of primate immunodeficiency viruses are essential transactivators to switch from early to late phase in the viral replication cycle. Surprisingly, the Rev protein of HIV-1 is able to substitute those of HIV-2 and, as shown in here, of SIVmac239, but not vice versa. To understand the underlying mechanism of this incomplete functional reciprocity, we constructed a series of chimeric HIV-1/SIVmac239 Rev proteins and tested for transcomplementation efficacy on Rev-dependent indicator plasmids. In addition, we analyzed the prokaryotically expressed wild type and chimeric proteins for RNA-binding properties in a gel-shift assay in vitro. The functional defect of SIVmac239 on the HIV-1 Rev response element (RRE) is not due to a lack of binding or multimerization. In cotransfection experiments, SIVmac239 Rev and the chimeric proteins were tested for potential inhibitory effects on HIV-1 Rev function using the HIV-1 based indicator plasmid. Some of these proteins turned out to be transdominant inhibitors almost as potent as the HIV-1 Rev mutant M10 which is localized in the activation domain and is one of the strongest transdominant inhibitors. Surprisingly, M10 was not able to inhibit the function of either Rev protein on SIVmac239 RRE, whereas a corresponding SIVmac239 Rev mutant (SIV M10) was a transdominant inhibitor of SIVmac239 Rev function on its homologous RRE.

Amino Acid Sequence↗

PubMed1992

Role of human T-cell leukemia virus type 1 X region proteins in immortalization of primary human lymphocytes in culture.

Human T-cell leukemia virus type 1 (HTLV-1) immortalizes human CD4+ T lymphocytes in culture. Previous studies show that in the context of a herpesvirus saimiri vector, the sequence of the X region at the 3' end of the HTLV-1 genome is also capable of immortalizing CD4+ lymphocytes in the absence of HTLV-1 structural proteins. The X region of HTLV-1 encodes two trans-acting viral proteins, the 42-kDa Tax protein and the 27-kDa Rex protein. Infection of human cord blood cells with herpesvirus saimiri recombinants which contain HTLV-1 X region sequences defective for expression of tax, rex, or both tax and rex demonstrates that tax function is necessary and sufficient for immortalization of primary human CD4+ cord blood lymphocytes in culture in the context of the herpesvirus saimiri vector.

Base Sequence↗

PubMed1991

In vitro binding of human T-cell leukemia virus rex proteins to the rex-response element of viral transcripts.

Human T-cell leukemia virus (HTLV-I, HTLV-II) rex protein function is required for the cytoplasmic expression of incompletely spliced viral transcripts encoding structural proteins. The effect is mediated by a cis-acting rex-response element (RRX) which is located near the 3' end of all viral mRNAs. We show that rex polypeptides of HTLV-I and HTLV-II expressed in Escherichia coli are capable of specifically binding RRX-containing transcripts of both viruses in cell-free assays. Binding analyses with deletion variants of rex proteins revealed a domain with RNA-binding activity in the first 77 N-terminal amino acids. Removal of a basic peptide of 19 amino acids from the N terminus abrogated RNA binding, whereas a beta-galactosidase fusion protein containing this peptide bound to the RRX. These results suggest that direct binding of rex protein to the RRX is important for rex-mediated regulation of viral gene expression and that a short stretch of positively charged amino acids contributes to the specific binding of rex to its target RNA.

Carrier Proteins↗