HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Free Full Text (PDF) Free Supplemental Data All Versions of this Article: fj.06-7598comv1 21/14/4131    most recent Alert me when this article is cited Alert me if a correction is posted 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 Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Lai, T.-S. Articles by Greenberg, C. S. Search for Related Content PubMed PubMed Citation Articles by Lai, T.-S. Articles by Greenberg, C. S.

Identification of two GTP-independent alternatively spliced forms of tissue transglutaminase in human leukocytes, vascular smooth muscle, and endothelial cells

Thung-S. Lai^*, Yusha Liu^*, Weidong Li^* and Charles S. Greenberg^*,,1

Departments of
^* Medicine and

Pathology, Duke University Medical Center, Durham, North Carolina, USA

¹Correspondence: Box 2603, Duke University Medical Center, Durham, NC 27710, USA. E-mail: charles.greenberg{at}duke.edu

Tissue transglutaminase (tTG) is a multifunctional enzyme with transglutaminase crosslinking (TGase), GTP binding, and hydrolysis activities that play a role in many different disorders. We identified, characterized, and investigated the function and stability of two alternatively spliced forms of tTG using biochemical, cellular, and molecular biological approaches. Using a human aortic vascular smooth muscle cells (VSMC) cDNA library, we identified two cDNAs encoding C-terminal truncated forms, tTG_V1 and tTG_V2. tTG_V1,2 mRNAs were synthesized by a rare splicing event using alternate splice sites within exons 12 and 13 of the tTG gene, respectively. Quantitative PCR and immunoblotting demonstrated that there was unique expression and localization of tTG_V1,2 compared with tTG in human umbilical vein endothelial cells (HUVECs), VSMC, and leukocytes. The loss of C-terminal 52 amino acid residues (AAs) in tTG_V1,2 altered GTP binding, enhanced GTP hydrolysis, rendered the variants insensitive to GTP inhibition, and resulted in <10% residual Ca⁺²-dependent TGase activity. Transfection in HEK293 demonstrated a 28- and 5-fold reduction in the expression of tTG_V1 and tTG_V2, respectively, demonstrating that the C-terminal GTP-binding domain is important in stabilizing and promoting the half-life of tTG. The altered affinity for GTP allowed tTG_V1,2 to exhibit enhanced TGase activity when there is a transient increase in Ca⁺² levels. The abundance of tTG_V1,₂ and its distinct intracellular expression patterns in human vascular cells and leukocytes indicate these isoforms likely have unique physiological functions.—Lai, T. S., Liu, Y., Li, W., Greenberg, C. S. Identification of two GTP-independent alternatively spliced forms of tissue transglutaminase in human leukocytes, vascular smooth muscle and endothelial cells.

Key Words: neurodegenerative disease • aging • cell differentiation • extracellular matrix • vascular disease