This Article Full Text Full Text (PDF) All Versions of this Article: 148/12/5955    most recent Author Manuscript (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Loesch, K. Articles by Frank, S. J. Search for Related Content PubMed PubMed Citation Articles by Loesch, K. Articles by Frank, S. J.

Endoplasmic Reticulum-Associated Degradation of Growth Hormone Receptor in Janus Kinase 2-Deficient Cells

Department of Cell Biology (K.L., L.D., S.J.F.) and Department of Medicine (X.W., K.H., J.J., S.J.F.), Division of Endocrinology, Diabetes, and Metabolism, University of Alabama at Birmingham, Birmingham, Alabama 35294; and Endocrinology Section (S.J.F.), Medical Service, Veterans Affairs Medical Center, Birmingham, Alabama 35233

Address all correspondence and requests for reprints to: Stuart J. Frank, University of Alabama at Birmingham, 1530 3rd Avenue South, BDB 861, Birmingham, Alabama 35294-0012. E-mail: sjfrank{at}uab.edu.

A key factor governing cellular sensitivity to GH is cell surface GH receptor (GHR) abundance, which is affected transcriptionally and posttranscriptionally. Mature cell surface GHR abundance is regulated by constitutive and inducible metalloproteolysis and constitutive endosomal/lysosomal degradation. We previously found that Janus kinase 2 (JAK2)-deficient GHR-expressing cells have a greater precursor/mature GHR ratio, exhibit diminished inducible metalloproteolysis, and have a cytoplasmic domain-containing GHR fragment called the basal remnant (by virtue of comigration on SDS-PAGE with the inducible, metalloprotease-generated remnant). Herein we examined the mechanism of generation of basal remnant in JAK2-deficient cells, asking whether it originates from precursor vs. mature receptor and which protease(s) catalyzes its appearance. Prolonged metalloprotease inhibitor treatment or small interfering RNA knockdown of TNF- converting enzyme (TACE) and a disintegrin and metalloprotease-10 (ADAM10) (both implicated in inducible GHR proteolysis) did not reduce basal remnant, indicating its generation is not metalloprotease dependent. However, a mutant GHR resistant to metalloprotease cleavage did not yield basal remnant when expressed in JAK2-deficient cells, suggesting common structural determinants for generation of the inducible remnant and the non-metalloprotease-generated basal remnant seen in JAK2-deficient cells. Treatment of JAK2-deficient cells with a proteasome inhibitor, but not two separate lysosome inhibitors, dramatically decreased basal remnant, accompanied by decreased precursor GHR and increased mature GHR abundance. Disruption of endoplasmic reticulum-to-Golgi transport with brefeldin A (BFA) also reduced basal remnant, and washout of BFA allowed regeneration of basal remnant along with GHR precursor. Notably, BFA washout in the presence of cycloheximide blocked both basal remnant and precursor GHR reappearance, but BFA washout in the presence of lactacystin blocked only basal remnant reappearance, suggesting that basal remnant is generated proteasome dependently from precursor GHR. Collectively, our data suggest that JAK2, by association with GHR in the secretory pathway, blunts proteasome activity-dependent discrete GHR cleavage and endoplasmic reticulum-dependent degradation of the precursor receptor. In so doing, JAK2 enables efficient processing of precursor receptor to mature GHR.