CRP2 is an autonomous actin‐binding protein

FEBS Letters

Grubinger, Markus¹  Gimona, Mario¹ 

[1] Institute of Molecular Biology, Department of Cell Biology, Austrian Academy of Sciences, Billrothstrasse 11, A-5020 Salzburg, Austria

关键词: Cysteine-rich protein;    Actin binding;    Green fluorescent protein;    α-Actinin;    Zyxin;    Cytoskeleton;    GFP;    green fluorescent protein;    CRP;    cysteine-rich protein;   

DOI  :  10.1016/S0014-5793(03)01451-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Cysteine-rich proteins (CRPs) have been shown to be involved in cell differentiation, transcriptional regulation and the organisation of the actin cytoskeleton. Thus far, the latter function has been inferred solely from the in vitro interaction of CRP1, CRP2, and CRP3 with α-actinin and zyxin. We show here that purified, recombinant CRP2 binds directly to F-actin in vitro in co-sedimentation assays. Using a green fluorescent protein (GFP)-tagged construct of CRP2 we analysed its localisation and dynamics in A7r5 rat smooth muscle cells. CRP2 was associated with the actin cytoskeleton and decorated actin stress fibres in a continuous fashion, unlike the periodic labelling pattern observed for α-actinin and zyxin, which also accumulate in focal adhesions. Using live video fluorescence microscopy we observed the behaviour of GFP-CRP2 during the dynamic rearrangement of the actin cytoskeleton in phorbol 12,13-dibutyrate-treated A7r5 cells. In contrast to the actin-binding proteins SM22α and α-actinin, GFP-CRP2 did not translocate into the podosomes induced by this treatment, but remained preferentially bound to the stress fibres, suggesting an actin filament-stabilising role for CRP2. When fused to the mitochondrial targeting sequence from the Listeria protein ActA, GFP-CRP2 was almost completely localised to mitochondria, but no significant recruitment of either α-actinin or zyxin could be observed. Taken together, our results demonstrate that CRP2 can bind to F-actin directly and that the association with the actin cytoskeleton is independent of α-actinin or zyxin localisation in the cell, thus questioning the role of CRP2 as a regulator of α-actinin function in vivo.

【 授权许可】

Unknown