A Transglutaminase Immunologically Related to Tissue Transglutaminase Catalyzes Cross-Linking of Cell Wall Proteins inChlamydomonas reinhardtii  

Abstract

The addition of primary amines to the growth medium of the unicellular green alga Chlamydomonas reinhardtii disrupts cell wall assembly in both vegetative and zygotic cells. Primary amines are competitive inhibitors of the protein-cross-linking activity of transglutaminases. Two independent assays for transglutaminase confirmed a burst of extracellular activity during the early stages of cell wall formation in both vegetative cells and zygotes. When noninhibiting levels of a radioactive primary amine ( 14 C-putrescine) were added to the growth medium, both cell types were labeled in a reaction catalyzed by extracellular transglutaminase. The radioactive label was found specifically in the cell wall proteins of both cell types, and acid hydrolysis of the labeled material released unmodified 14 C-putrescine. Western blots of the proteins secreted at the times of maximal transglutaminase activity in both cell types revealed a single highly cross-reactive 72-kD band when screened with antibodies to guinea pig tissue transglutaminase. Furthermore, the proteins immunoprecipitated by this antiserum in vivo exhibited transglutaminase activity. We propose that this transglutaminase is responsible for an early cell wall protein cross-linking event that temporally precedes the oxidative cross-linking mediated by extracellular peroxidases.

14-3-3 Proteins Are Constituents of the Insoluble Glycoprotein Framework of the Chlamydomonas Cell Wall

“…Molecular genetic evidence indicates that V. carteri evolved from a unicellular ancestor similar to Chlamydomonas during the past 50 million years (Rausch et al, 1989). Two mechanisms for the cross-linking of cell wall glycoproteins have been described for Chlamydomonas: isodityrosine formation by peroxidase-catalyzed cross-linking of Tyr residues (Waffenschmidt et al, 1993) and cross-linking by transglutaminase (Waffenschmidt et al, 1999). Simultaneous binding of two soluble cell wall glycoproteins to one 14-3-3 molecule could facilitate their cross-linking by peroxidase or transglutaminase or an autocatalytic reaction.…”

“…parent molecular masses of 25 and 33 kD, which correspond to those of free 14-3-3 subunits, whereas the 14-3-3 polypeptides released from the insoluble wall fraction by treatment with HF/pyridine are considerably larger (45 and 64 kD; Figure 4A). Therefore, we assume that only a small portion of the 14-3-3 proteins that occur in the cell wall are linked covalently to the insoluble cell wall fraction by peroxidase-catalyzed isodityrosine formation and/or transglutaminase activity (Waffenschmidt et al, 1993(Waffenschmidt et al, , 1999, whereas the predominant portion remains soluble and is accumulated subsequently in the culture medium.…”

“…Isodityrosine has been detected in hydrolysates of Chlamydomonas cell walls (Waffenschmidt et al, 1993), indicating that peroxidase-catalyzed cross-linking of cell wall glycoproteins via Tyr side chains, which has been observed for the extensins of higher plants (Fry, 1982;Epstein and Lamport, 1984;Biggs and Fry, 1990), also occurs in Chlamydomonas. Furthermore, it has been reported that a transglutaminase also catalyzes the cross-linking of cell wall proteins in Chlamydomonas (Waffenschmidt et al, 1999).…”

Transglutaminase activity during senescence and programmed cell death in the corolla of tobacco (Nicotiana tabacum) flowers

“…25 Evidence for their structural role comes from the identification of some of their substrates, e.g. actin and tubulin 24 or cell wall proteins, 21 and through its involvement in the polymerisation of Rubisco. 26 Different roles for TGase have been proposed in chloroplasts 27 and mitochondria.…”

Sulfur Economy and Cell Wall Biosynthesis during Sulfur Limitation of Chlamydomonas reinhardtii

“…The cell wall HRGPs form a crystal structure with a central trilaminar morphology embedded in a meshwork of fibrils (Hills et al, 1975;Goodenough and Heuser, 1985;Goodenough et al, 1986;Adair et al, 1987). Crosslinking among the components of the wall may occur through the formation of isodityrosine bridges (Waffenschmidt et al, 1993) and as a consequence of transglutaminase activity (Waffenschmidt et al, 1999). In this study, we identified two extracellular polypeptides, Ecp76 and Ecp88, that specifically accumulate when C. reinhardtii cells are starved for sulfur.…”