Interaction of microtubules and actin with the N-terminus of βPix-b(L) directs cellular pinocytosis.
βPix is a Rac/Cdc42 guanine nucleotide exchange factor (GEF) that is known to be a regulator of actin cytoskeleton remodeling. Recently, a novel splicing isoform, βPix-b(L), was identified as an alternative translational product of the βPix-b mRNA with an extended N-terminus comprising a partial calponin homology (CH) domain and a serine-rich (SR) domain. However, the cellular function of βPix-b(L) is largely unknown. In the current study, we analyzed the genomic DNA structure and cellular functions of βPix-b(L). The results of this study demonstrate that βPix is composed of 24 exons and 21 introns spanning around 100 kb. RT-PCR experiments revealed that there are two forms of βPix mRNA with distinct 5' UTRs that are the result of alternative splicing of exon 1 and 2 from βPix genomic DNA. In addition, affinity chromatography analysis and a pull-down assay with the N-terminal region of βPix-b(L) revealed that βPix-b(L) interacts with tubulin and actin via its N-terminal CH and SR domains, respectively. Interaction with tubulin enabled βPix-b(L) to bundle the microtubule and form membrane protrusions. Furthermore, the N-terminus of βPix-b(L) was also critical for its localization to cellular vesicles. Functionally, βPix-b(L) induced pinocytosis through cooperative action of the CH and Dbl homology (DH) domains, demonstrating the role of βPix-b(L) in the regulation of membrane dynamics.
School of Biological Sciences, Seoul National University, Seoul 151-742, Korea.
Seung Joon
SJ
Lee
Soo Jung
SJ
Yang
Dae Hwan
DH
Kim
Jhang Ho
JH
Pak
Kwang Ho
KH
Lee
Kyoung Hee
KH
Choi
Dongeun
D
Park
Sangmyung
S
Rhee
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity; D017931:DNA Primers; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D007150:Immunohistochemistry; D051379:Mice; D008870:Microtubules / Q000378:metabolism*; D010873:Pinocytosis; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D064067:Rho Guanine Nucleotide Exchange Factors
C576607:Arhgef7 protein, mouse; D017931:DNA Primers; D020662:Guanine Nucleotide Exchange Factors; D012333:RNA, Messenger; D064067:Rho Guanine Nucleotide Exchange Factors
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