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PMID: 1310321 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Targeted disruption of the ABP-120 gene leads to cells with altered motility.

The Journal of cell biology ·Vol. 116 ·No. 4 ·1992-02-00 ·Pages 943-55

Cox D, Condeelis J, Wessels D, Soll D, Kern H, Knecht DA

Abstract

The actin-binding protein ABP-120 has been proposed to play a role in cross-linking F-actin filaments during pseudopod formation in motile Dictyostelium amebas. We have tested this hypothesis by analyzing the phenotype of mutant cell lines which do not produce ABP-120. Two different transformation vectors capable of targeted disruption of the ABP-120 gene locus have been constructed using a portion of an ABP-120 cDNA clone. Three independent cell lines with different disruption events have been obtained after transformation of amebas with these vectors. The disruption of the ABP-120 gene by vector sequences results in either the production of a small amount of truncated ABP-120 or no detectable protein at all. The phenotypes of two different clones lacking ABP-120, generated in strains AX3 and AX4, have been characterized and show identical results. ABP-120- cells tend to remain rounder before and after cAMP stimulation, and do not reextend pseudopods normally after rapid addition of cAMP. In addition, ABP-120- cells translocating in buffer exhibit defects in both the rate and extent of pseudopod formation. The amount of F-actin cross-linked into the cytoskeleton after cAMP stimulation of ABP-120- cells is reduced at times when ABP-120 has been shown to be incorporated into the cytoskeleton, and this correlates temporally with the absence of reextension of pseudopods after cAMP stimulation. The instantaneous velocity is significantly reduced both before and after cAMP stimulation in the ABP-120- cells, and the cells show decreased chemotactic efficiency compared to ABP-120+ controls. This phenotype is consistent with a role for ABP-120 in pseudopod extension by cross-linking actin filaments as proposed by the "cortical expansion model" (Condeelis, J., A. Bresnick, M. Demma, C. Dharmawardhane, R. Eddy, A. L. Hall, R. Sauterer, and V. Warren. 1990. Dev. Genet. 11:333-340).

MeSH Terms
Actins/metabolism Animals Blotting, Northern Carrier Proteins/genetics,physiology Cell Line Cell Movement/drug effects Chemotaxis Cyclic AMP/pharmacology Cytoskeleton/metabolism Dictyostelium/genetics,physiology Genetic Vectors Microfilament Proteins/genetics,physiology Mutation Phenotype Pseudopodia/physiology,ultrastructure Transformation, Genetic
Chemicals
Actins Carrier Proteins Microfilament Proteins abpC protein, Dictyostelium Cyclic AMP
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Cox D
Albert Einstein College of Medicine, Yeshiva University, Bronx, New York.
Condeelis J
Wessels D
Soll D
Kern H
Knecht D A
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.116.4.943
Published
1992-02-00
Pages
943-55
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2289347
Subset
IM
Grants
NHLBI NIH HHS · 5T32 HL07675-03 · United States
NICHD NIH HHS · HD07216 · United States
NICHD NIH HHS · HD18577 · United States
External Links
PubMed source DOI: 10.1083/jcb.116.4.943 PMC Full Text View journal details
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