title: Tissue transglutaminase promotes serotonin-induced AKT signaling and mitogenesis in pulmonary vascular smooth muscle cells.
abstract: Tissue transglutaminase 2 (TG2) is a multifunctional enzyme that cross-links proteins with monoamines such as serotonin (5-hydroxytryptamine, 5-HT) via a transglutamidation reaction, and is associated with pathophysiologic vascular responses. 5-HT is a mitogen for pulmonary artery smooth muscle cells (PASMCs) that has been linked to pulmonary vascular remodeling underlying pulmonary hypertension development. We previously reported that 5-HT-induced PASMC proliferation is inhibited by the TG2 inhibitor monodansylcadaverine (MDC); however, the mechanisms are poorly understood. In the present study we hypothesized that TG2 contributes to 5-HT-induced signaling pathways of PASMCs. Pre-treatment of bovine distal PASMCs with varying concentrations of the inhibitor MDC led to differential inhibition of 5-HT-stimulated AKT and ROCK activation, while p-P38 was unaffected. Concentration response studies showed significant inhibition of AKT activation at 50 μM MDC, along with inhibition of the AKT downstream targets mTOR, p-S6 kinase and p-S6. Furthermore, TG2 depletion by siRNA led to reduced 5-HT-induced AKT activation. Immunoprecipitation studies showed that 5-HT treatment led to increased levels of serotonylated AKT and increased TG2-AKT complex formations which were inhibited by MDC. Overexpression of TG2 point mutant cDNAs in PASMCs showed that the TG2 C277V transamidation mutant blunted 5-HT-induced AKT activation and 5-HT-induced PASMC mitogenesis. Finally, 5-HT-induced AKT activation was blunted in SERT genetic knock-out rat cells, but not in their wild-type counterpart. The SERT inhibitor imipramine similarly blocked AKT activation. These results indicate that TG2 contributes to 5-HT-induced distal PASMC proliferation via promotion of AKT signaling, likely via its serotonylation. Taken together, these results provide new insight into how TG2 may participate in vascular smooth muscle remodeling.
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Krishna
authors.initials: K
authors.lastname: Penumatsa
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Shereen
authors.initials: S
authors.lastname: Abualkhair
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Lin
authors.initials: L
authors.lastname: Wei
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Rod
authors.initials: R
authors.lastname: Warburton
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Ioana
authors.initials: I
authors.lastname: Preston
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Nicholas S
authors.initials: NS
authors.lastname: Hill
authors.affiliation: Dept of Pharmacology and Toxicology, Michigan State University, East Lansing, MI 48824, United States.
authors.forename: Stephanie W
authors.initials: SW
authors.lastname: Watts
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States.
authors.forename: Barry L
authors.initials: BL
authors.lastname: Fanburg
authors.affiliation: Pulmonary and Critical Care Division, Tupper Research Institute, Tufts Medical Center, Tufts University School of Medicine, Boston, MA 02111, United States. Electronic address: dtoksoz@tuftsmedicalcenter.org.
authors.forename: Deniz
authors.initials: D
authors.lastname: Toksoz
mesh_terms: D000818:Animals; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D002417:Cattle; D018076:DNA, Complementary / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D007099:Imipramine / Q000494:pharmacology; D008938:Mitosis / Q000187:drug effects*; D050505:Mutant Proteins / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D011485:Protein Binding / Q000187:drug effects; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011651:Pulmonary Artery / Q000166:cytology*; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D038601:Ribosomal Protein S6 / Q000378:metabolism; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism; D012701:Serotonin / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D011503:Transglutaminases / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D050505:Mutant Proteins; D034741:RNA, Small Interfering; D038601:Ribosomal Protein S6; C000717609:Tgm2 protein, rat; D012701:Serotonin; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D051057:Proto-Oncogene Proteins c-akt; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D054460:rho-Associated Kinases; D019204:GTP-Binding Proteins; C008542:monodansylcadaverine; D002103:Cadaverine; D007099:Imipramine; D013936:Thymidine
keywords: AKT; S6; S6K; SERT; Serotonin; Transglutaminase 2