Publications
Department of Medicine faculty members published more than 3,000 peer-reviewed articles in 2022.
1994
1994
1994
1994
Laminin is a heterotrimeric glycoprotein found in the perisinusoidal space of adult rat liver. The principal cellular source of laminin in liver is the lipocyte, with its three subunits measuring 324, 200 and 200 kD. The large subunit of lipocyte-derived laminin is distinct from the A subunit of murine laminin (440 kD); its size suggests that it represents a peptide, called M, recently cloned from human placenta. Using oligonucleotide primers derived from the human M-subunit cDNA, we amplified a 445-bp sequence encoding a fragment of M-laminin from adult rat lipocytes. The rat cDNA is 90% homologous to the human M-subunit cDNA and recognizes an mRNA in lipocytes measuring about 10 kb. M-subunit transcripts were identified only in lipocytes from normal adult liver; they could not be identified in hepatocytes, endothelial cells or Kupffer cells. Lipocytes were screened for M-subunit protein with a polyclonal M antiserum. Cells stained specifically for the M-subunit after 36 hr in primary culture; the protein was also identified in freshly isolated cells by means of immunoblotting. To determine whether lipocytes alter their expression of the laminin M subunit during liver injury, we monitored M-subunit mRNA in these cells at various intervals after carbon tetrachloride administration. M-subunit transcripts increased twofold within 12 hr of toxin exposure, returning to below baseline by 48 hr. The results indicate that lipocytes produce the M subunit of laminin in place of A. Production of this subunit by lipocytes may facilitate cell growth and reorganization during liver regeneration.
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1994
1994
It is now widely appreciated that G-protein-coupled cell-surface receptors can modulate distinct signal transduction pathways via coupling to different GTP-binding proteins. In the present study, we have used a transient co-expression approach to study the coupling of a single alpha 2-adrenergic receptor (alpha 2AAR) population to three different G protein subtypes (Gi, Gq, and Gs) acting on two different cellular effectors in HEK 293 cells. In all cases, the affinity of the receptor for the alpha 2A-adrenergic agonist, UK14304, is unchanged (KD approximately equal to 670 nM). However, there is a dramatic difference in the EC50 of UK14304 in eliciting inhibition of endogenous adenylyl cyclase via endogenous Gi (0.09 nM) versus activation of phospholipase C via co-transfected Gq (50 nM) or stimulation of endogenous adenylyl cyclase via co-transfected Gs (70 nM) in HEK 293 cells. These findings are consistent with the interpretations that the alpha 2AAR preferentially interacts with Gi rather than Gs or Gq. When the alpha 2AAR was mutated at Asp79, a residue highly conserved among G-protein-coupled receptors, the mutant D79N alpha 2AAR lost the ability to couple to Gq and Gs and, although it was able to couple to inhibition of cyclase via pertussis toxin-sensitive pathways (Gi), it did so with a lower potency than observed for the wild-type alpha 2AAR (EC50 = 7.2 nM). The most straightforward interpretation of these data is that the D79N mutation in the alpha 2AAR reduces the efficiency of coupling of the alpha 2AAR to all G-proteins, thus eliminating signal transduction through those pathways less efficiently coupled to the alpha 2AAR. Since the transient expression assays described permit manipulation of the structure of both the receptor or the G-protein, the present strategies could be exploited to delineate the complementary domains specifying the affinity and/or efficacy of receptor coupling to distinct GTP-binding proteins.
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