β-Thymosins are not simple actin monomer buffering proteins

Insights from overexpression studies

Hui Qiao Sun, Katarzyna Kwiatkowska, Helen Lu Yin

Research output: Contribution to journalArticle

52 Citations (Scopus)

Abstract

β-Thymosins are the currently favored candidates for maintaining the large actin monomer (G-actin) pool in living cells. To determine if β-thymosin behaves like a simple G-actin buffering agent in the complex environment of a cell, we overexpressed thymosin β10 (Tβ10) in NIH3T3 cells and determined the effect on the monomer/ polymer equilibrium. Tβ10 is the predominant β-thymosin isoform in the NIH3T3 cell line, and it is present in approximately equal molar ratio to profilin and cofilin/ actin depolymerizing factor, two other well characterized actin monomer binding proteins. Clonal cell lines that overexpressed three times more Tβ10 had 23-33% more polymerized actin than control cells, and the filaments appeared thicker after staining with fluorescent phalloidin. There was no change in total actin, profilin, and cofilin/actin depolymerizing factor content. The overexpressing cells were more motile; they spread faster and had higher chemotactic and wound healing activity. Assuming that there is no compensatory inactivation of the other classes of monomer binding proteins, our paradoxical observation can be accounted for quantitatively by a parallel in vitro study (Carlier, M.-F., Didry, D., Erk, I., Lepault, J., Van Troys, L., Vanderkekove, J., Perelroizen, I., Yin, H. L., Doi, Y., and Pantaloni, D., (1996) J. Biol. Chem. 271, 9231-9239). β-Thymosin at levels comparable with that found in the overexpressing cells binds actin filaments and decreases the critical concentration (Cc) for actin polymerization. This reduces the monomer buffering ability of ß-thymosin, so that above a certain threshold an incremental increase in thymosin does not lead to a corresponding increase in G-actin. Furthermore, the decrease in Cc reduces the buffering capacity of the other actin monomer binding proteins. As a consequence, an increase in β-thymosin does not necessarily result in a proportionate increase in actin monomer content in a complex environment containing other actin monomer binding proteins. The outcome depends on the level of β-thymosin expression relative to the composition of the other actin monomer binding protien. Our results suggest that β-thymosins are not simple actin buffering proteins and that their biphasic action may have physiological significance.

Original languageEnglish (US)
Pages (from-to)9223-9230
Number of pages8
JournalJournal of Biological Chemistry
Volume271
Issue number16
StatePublished - Apr 19 1996

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Thymosin
Actins
Monomers
Proteins
Microfilament Proteins
Destrin
Profilins
Actin Depolymerizing Factors
Carrier Proteins
Cells
Phalloidine
Cell Line
Actin Cytoskeleton
Polymerization
Wound Healing
Polymers

ASJC Scopus subject areas

  • Biochemistry

Cite this

β-Thymosins are not simple actin monomer buffering proteins : Insights from overexpression studies. / Sun, Hui Qiao; Kwiatkowska, Katarzyna; Yin, Helen Lu.

In: Journal of Biological Chemistry, Vol. 271, No. 16, 19.04.1996, p. 9223-9230.

Research output: Contribution to journalArticle

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abstract = "β-Thymosins are the currently favored candidates for maintaining the large actin monomer (G-actin) pool in living cells. To determine if β-thymosin behaves like a simple G-actin buffering agent in the complex environment of a cell, we overexpressed thymosin β10 (Tβ10) in NIH3T3 cells and determined the effect on the monomer/ polymer equilibrium. Tβ10 is the predominant β-thymosin isoform in the NIH3T3 cell line, and it is present in approximately equal molar ratio to profilin and cofilin/ actin depolymerizing factor, two other well characterized actin monomer binding proteins. Clonal cell lines that overexpressed three times more Tβ10 had 23-33{\%} more polymerized actin than control cells, and the filaments appeared thicker after staining with fluorescent phalloidin. There was no change in total actin, profilin, and cofilin/actin depolymerizing factor content. The overexpressing cells were more motile; they spread faster and had higher chemotactic and wound healing activity. Assuming that there is no compensatory inactivation of the other classes of monomer binding proteins, our paradoxical observation can be accounted for quantitatively by a parallel in vitro study (Carlier, M.-F., Didry, D., Erk, I., Lepault, J., Van Troys, L., Vanderkekove, J., Perelroizen, I., Yin, H. L., Doi, Y., and Pantaloni, D., (1996) J. Biol. Chem. 271, 9231-9239). β-Thymosin at levels comparable with that found in the overexpressing cells binds actin filaments and decreases the critical concentration (Cc) for actin polymerization. This reduces the monomer buffering ability of {\ss}-thymosin, so that above a certain threshold an incremental increase in thymosin does not lead to a corresponding increase in G-actin. Furthermore, the decrease in Cc reduces the buffering capacity of the other actin monomer binding proteins. As a consequence, an increase in β-thymosin does not necessarily result in a proportionate increase in actin monomer content in a complex environment containing other actin monomer binding proteins. The outcome depends on the level of β-thymosin expression relative to the composition of the other actin monomer binding protien. Our results suggest that β-thymosins are not simple actin buffering proteins and that their biphasic action may have physiological significance.",
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