TRAM2 protein interacts with endoplasmic reticulum Ca2+ pump Serca2b and is necessary for collagen type I synthesis.
Cotranslational insertion of type I collagen chains into the lumen of the endoplasmic reticulum (ER) and their subsequent folding into a heterotrimeric helix is a complex process which requires coordinated action of the translation machinery, components of translocons, molecular chaperones, and modifying enzymes. Here we describe a role for the protein TRAM2 in collagen type I expression in hepatic stellate cells (HSCs) and fibroblasts. Activated HSCs are collagen-producing cells in the fibrotic liver. Quiescent HSCs produce trace amounts of type I collagen, while upon activation collagen synthesis increases 50- to 70-fold. Likewise, expression of TRAM2 dramatically increases in activated HSCs. TRAM2 shares 53% amino acid identity with the protein TRAM, which is a component of the translocon. However, TRAM2 has a C terminus with only a 15% identity. The C-terminal part of TRAM2 interacts with the Ca(2+) pump of the ER, SERCA2b, as demonstrated in a Saccharomyces cerevisiae two-hybrid screen and by immunoprecipitations in human cells. TRAM2 also coprecipitates with anticollagen antibody, suggesting that these two proteins interact. Deletion of the C-terminal part of TRAM2 inhibits type I collagen synthesis during activation of HSCs. The pharmacological inhibitor of SERCA2b, thapsigargin, has a similar effect. Depletion of ER Ca(2+) with thapsigargin results in inhibition of triple helical collagen folding and increased intracellular degradation. We propose that TRAM2, as a part of the translocon, is required for the biosynthesis of type I collagen by coupling the activity of SERCA2b with the activity of the translocon. This coupling may increase the local Ca(2+) concentration at the site of collagen synthesis, and a high Ca(2+) concentration may be necessary for the function of molecular chaperones involved in collagen folding.
Stefanovic
Branko
B
Department of Biomedical Science, College of Medicine, Florida State University, Tallahassee, Florida 32306, USA. branko.stefanovic@med.fsu.edu
Stefanovic
Lela
L
Schnabl
Bernd
B
Bataller
Ramon
R
Brenner
David A
DA
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D017384:Sequence Deletion; D019284:Thapsigargin / Q000494:pharmacology
D024042:Collagen Type I; D004220:Disulfides; D007093:Imidazoles; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; C486173:TRAM2 protein, human; C074282:translocating chain-associating membrane protein (TRAM); C031938:calmidazolium; D019284:Thapsigargin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium