Biomedical subjects

R Y Liu

Publications and source records attributed to R Y Liu.

31 records · Page 2Linked to original sources

PubMed1992

Expression of human hsp70 in rat fibroblasts enhances cell survival and facilitates recovery from translational and transcriptional inhibition following heat shock.

We report here our studies on the inhibitional effect of 45 degrees C heat shock on transcriptional and translational activity and their subsequent recovery at 37 degrees C in Rat-1 cells, thermotolerant Rat-1 cells (TT Rat-1), and Rat-1 cells transfected with human hsp70 gene (HR24, M21). Specifically, we ask whether overexpression of hsp70 protects cells from heat-induced inhibition in RNA and protein synthesis, and/or facilitates cells' ability to recover from translational or transcriptional inhibition after heat shock treatment. Our data demonstrate that the constitutive expression of human hsp70, by itself, confers thermal resistance as expressed in enhanced survival and translational tolerance, but not transcriptional tolerance. In addition, the expression of human hsp70 in Rat-1 cells facilitates cells' ability to recover from heat-induced inhibition in protein and RNA synthesis. After healing at 45 degrees C for 25 min, the time required for RNA and protein synthesis to recover is considerably shorter in HR24, M21, and TT Rat-1 cells than that in control Rat-1 cells. These results provide strong evidence for a direct link between the expression of a functional form of mammalian hsp70, and cells' translational tolerance, as well as their ability to recover from translational and transcriptional inhibition after heat shock.

Animals↗

PubMed1992

Heat shock protein hsp70 protects cells from thermal stress even after deletion of its ATP-binding domain.

Retroviral-mediated gene transfer experiments show that rodent cells become heat resistant when stably and constitutively expressing a cloned human gene encoding an intact human 70-kDa heat shock protein (hsp70). Cells expressing higher levels of the hsp70 protein generally tolerate thermal stress better, whereas cells expressing either of two mutated hsp70-encoding genes, one with a 4-base pair out-of-frame deletion and one with an in-frame deletion of codons 438-618, are heat sensitive. These results provide strong evidence that expression of hsp70 leads directly to thermal tolerance. Surprisingly, cells expressing a mutant hsp70 of a human gene missing codons 120-428 are, nevertheless, heat resistant. Because the deleted region of this mutant contains the ATP-binding domain of human hsp70, this domain appears dispensable in the hsp70-mediated protection of cells from thermal stress.

Adenosine Triphosphate↗

PubMed1992

Identification of human hsp-70 expression in stably transfected rodent cells by isoelectrofocus immunoblots.

The major heat-shock protein, hsp-70, is synthesized by cells from a wide variety of organisms in response to heat shock or other stresses. It is assumed that hsp-70 may have an important thermal protective function. To test this hypothesis directly, we have transfected rat fibroblast cells with appropriate expression plasmids containing a cloned human hsp-70 gene. Stable transfectants expressing the human hsp-70 gene product were identified by Western blot analysis. During the course of selecting successful transfectants, we found that when standard methods of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunostaining were employed with the monoclonal antibody raised against the human hsp-70 antigen, we were unable to differentiate the human hsp-70 from the heat-inducible rat hsp-70. This was because the monoclonal antibody cross-reacts with the human and rat proteins, which have the same mobility in SDS-PAGE, and it is difficult to determine which protein is expressed. To improve the resolution of the Western blot technique, we performed additional immunoblot analysis of cellular proteins separated by slab gel isoelectrofocusing. Our study shows that the isoelectrofocusing technique, when combined with antibodies against hsp-70, gave a better resolution for the separation of exogenous human hsp-70 and the endogenous rat hsp-70 than the commonly used SDS-PAGE Western blot analysis. It provides a rapid and specific method to identify positive colonies that express the human hsp-70 gene in transfected rodent cells.

Animals↗

PubMed1990

[Facilitation of arginine-vasopressin analogs on learning and memory in rats].

Adult rats were trained in a maze for brightness discrimination (BD) or in a shuttle box for active avoidance. It was found that the pretreatment of 1-desamino-8-D-arginine-vasopressin (DDAVP) enhanced the retention of brightness discrimination and retarted the extinction of avoidance response. Neonatal daily administration for 14 d of both 9-desglycylamide-DDAVP (DGDDAVP) or 2-destyrosyl-3-desphenylalanyl-9-desglycylamide-AVP (AVP4-8), but not D-arginine-AVP4-8 (DAVP4-8) showed significant facilitation on acquisition and subsequent maintenance of BD in adult rats. The results indicate that some of synthetic arginine-vasopressin analogs may directly act on CNS and consequently affect learning behaviors in rats.

Animals↗

PubMed1989

Expression of human placental aromatase in Saccharomyces cerevisiae.

A full-length human placental aromatase cDNA clone, Aro 2, was isolated upon screening a human placental cDNA library with an aromatase cDNA probe and an oligonucleotide probe whose sequence was derived from a human aromatase genomic clone. Nucleotide sequence microheterogeneity was found in the 3'-untranslated region among Aro 2 and in two previously described human aromatase cDNA clones. Both the minor sequence differences and the expression of a single protein species in placental tissue suggest the presence of different alleles for aromatase. Northern blot analyses using one cDNA and two oligonucleotide probes are consistent with the two mRNA messages of 2.9 and 2.5 kilobases arising in human placenta as a consequence of differential processing. Several yeast expression plasmids containing the aromatase cDNA we cloned were constructed. The enzyme was expressed in Saccharomyces cerevisiae. The expressed activity was inhibited by the known aromatase inhibitor, 4-hydroxyandrostenedione. A level of 2 micrograms aromatase/mg partially purified yeast microsomes was estimated by analyses of carbon monoxide difference spectra on microsomal fractions from yeast carrying plasmid pHARK/VGAL. Using [1 beta, 2 beta-3H]androst-4-ene-3,17-dione as the substrate, an apparent Michaels-Menken constant (Km) of 34 nM and a maximum velocity (Vmax) of 23 pmol [3H]water formed per min/mg protein were obtained for the yeast synthesized aromatase by transformation with plasmid pHARK/VGAL. The kinetic results are similar to those determined for human placental aromatase, and suggest that the yeast synthesized aromatase will be useful for further structure-function studies.

Amino Acid Sequence↗

PubMed1988

On a notion of simplicial depth.

For a distribution F on R(p) and a point x in R(p) the simplicial depth D(x), which is the probability that x be inside a random simplex whose vertices are p + 1 independent observations from F, is introduced. D(x) can be viewed as a measure of depth of the point x relative to F, and its empirical version gives rise to a natural ordering of the data points from the center outward. This ordering provides an approach for detecting outliers in a multivariate data cloud and leads to the introduction of affine equivariant multivariate generalizations of the univariate sample median and L-statistics. This sample median is shown to be consistent for the center of any angularly symmetric distribution.

Journal Article↗

PubMed1988