Expression pattern of SVEP1 alternatively-spliced forms.
SVEP1 is a multi-domain protein recognized as a cell adhesion molecule (CAM). In this study, we focused on the activity regulation of an alternative promoter (AP) and the expression of alternative splice forms of mRNA from SVEP1 gene. The expression of SVEP1 isoforms was analyzed on RNA isolated from pre-osteoblastic MBA-15 and mammary adenocarcinoma DA3 cells grown alone or following co-culture between these cells. The co-culture system aimed to mimic the cellular cross talk that exists in the bone microenvironment once the mammary cells invade the bone. We demonstrated that SVEP1 isoforms were differentially expressed between these cells. The various isoforms levels were affected by co-culturing or in cells treated with TNFα or estrogen. Both cell lines exhibited an increase of message levels of a and e isoforms following the co-culture conditions. A novel aspect presented here is related to existence of an alternative promoter (AP) in SVEP1 gene. The AP was in silico predicted and analyzed for binding by specific transcription factors (TFIIB, ERα, NF-κB, Sp1 and pcJUN) using Chromatin immunoprecipitation (ChIP) assay. The binding of these TFs results in a non uniform binding pattern when comparing between the DA3 and MBA-15 cells. Using the demethylation agent, 5'-aza-deoxycitidine and histone deacetylase inhibitor, Trichostatin-A allowed to study the methylation level of the AP and the message expression. This study provides insights into alternative splice forms of SVEP1 and their regulation that may play a role within the bone niche with invading carcinoma cells.
Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Ramat-Aviv, Israel.
Chen
C
Glait-Santar
Metsada
M
Pasmanik-Chor
Dafna
D
Benayahu
D017398:Alternative Splicing / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001374:Azacitidine / Q000494:pharmacology; D002135:Calcium-Binding Proteins; D015815:Cell Adhesion Molecules / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D018920:Coculture Techniques; D004967:Estrogens / Q000494:pharmacology; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D051379:Mice; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000964:Antimetabolites, Antineoplastic; D002135:Calcium-Binding Proteins; D015815:Cell Adhesion Molecules; D004967:Estrogens; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; C418635:Polydom protein, mouse; D011506:Proteins; C508516:SVEP1 protein, human; C549928:SVEP1 protein, rat; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; C012589:trichostatin A; D001374:Azacitidine