ANALYSIS OF NUCLEAR PROTEIN COMPLEXES COMPRISING α-ACTININ-4
BY 2D-ELECTROPHORESIS AND MASS-SPECTROMETRY
M. G. Khotin,1,* L. V. Turoverova,1 E. P. Podolskaya,2 I. A. Krasnov,2
A. V. Solovieva,2 V. Y. Aksenova,1 K.-E. Magnusson,3 G. P. Pinaev,1 D. G. Tentler 1
1 Institute of Cytology RAS, 2 Institute for Analytical Instrumentation RAS, St. Petersburg,
and 3 Linkoping University, Department of Clinical and Experimental Medicine, Sweden;
* e-mail: h_mg@mail.ru
Actin-binding protein α-actinin-4 is a member of spectrin super family. It is located in the cytoplasm and in
the nucleus. However, nuclear functions of α-actinin-4 are still not clear. In this study, we analyzed composition
of nuclear protein complexes associated with α-actinin-4 in A431 cells. Using 2D electrophoresis, we have
determined that about 50 different proteins may be associated with nuclear α-actinin-4. Using mass-spectrometry,
we analyzed major proteins of these complexes. β-Actin, α- and β-tubulins, ribonucleoprotein A2/B1, which
regulates splicing and is associated with β-actin, peroxiredoxin-1, which is involved in oxidative stress, and glycolytic
enzyme D-3-phosphoglycerate dehydrogenase were identified by MALDI-TOF. Detection of these proteins
in nuclear complexes with α-actinin-4 may suggest that α-actinin-4 is involved in transcription and splicing.
Presence of β-actin in the investigated complexes was confirmed by tandem mass-spectrometry (MALDITOF-
TOF). Immunoprecipitation of nuclear proteins with antibodies against α-tubulin confirmed association of
α-actinin-4 with α-tubulin in the protein complex. Nuclear α-actinin-4 constitutes of 105 KDa fullsize isoform
and two truncated isoforms of 65 and 75 kDa, whereas only the truncated isoform have been found in nuclear
complexes with α-tubulin. These data suggest that α-actinin-4 is associated with a number of different nuclear
protein complexes which may carry out different functions in the cell nucleus.
Key words: α-actinin-4, mass-spectrometry
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