Abstract
Twinfilin is a highly conserved member of the actin depolymerization
factor homology (ADF-H) protein superfamily, which also includes ADF/Cofilin,
Abp1/Drebrin, GMF, and Coactosin. Twinfilin has a unique molecular architecture
consisting of two ADF-H domains joined by a linker and followed by a C-terminal
tail. Yeast Twinfilin, in conjunction with yeast cyclase-associated protein
(Srv2/CAP), increases the rate of depolymerization at both the barbed and
pointed ends of actin filaments. However, it has remained unclear whether these
activities extend to Twinfilin homologs in other species. To address this, we
purified the three mouse Twinfilin isoforms (mTwf1, mTwf2a, mTwf2b) and mouse
CAP1, and used TIRF microscopy assays to study their effects on filament
disassembly. Our results show that all three mouse Twinfilin isoforms accelerate
barbed end depolymerization similar to yeast Twinfilin, suggesting that this
activity is evolutionarily conserved. In striking contrast, mouse Twinfilin
isoforms and CAP1 failed to induce rapid pointed end depolymerization. Using
chimeras, we show that the yeast-specific pointed end depolymerization activity
is specified by the C-terminal ADF-H domain of yeast Twinfilin. Additionally,
Tropomyosin decoration of filaments failed to impede depolymerization by yeast
and mouse Twinfilin and Srv2/CAP, but inhibited Cofilin severing. Together, our
results indicate that Twinfilin has conserved functions in regulating barbed end
dynamics, although its ability to drive rapid pointed end depolymerization
appears to be species-specific. We discuss the implications of this work,
including that pointed end depolymerization may be catalyzed by different ADF-H
family members in different species.