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Vertebrate isoforms of actin capping protein beta have distinct functions In vivo.

Hart MC, Cooper JA - J. Cell Biol. (1999)

Bottom Line: The beta2 did not localize to the Z-line.Therefore, CPbeta1 and CPbeta2 each have a distinct function that cannot be provided by the other isoform.CPbeta1 attaches actin filaments to the Z-line, and CPbeta2 organizes the actin at the intercalated discs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Actin capping protein (CP) binds barbed ends of actin filaments to regulate actin assembly. CP is an alpha/beta heterodimer. Vertebrates have conserved isoforms of each subunit. Muscle cells contain two beta isoforms. beta1 is at the Z-line; beta2 is at the intercalated disc and cell periphery in general. To investigate the functions of the isoforms, we replaced one isoform with another using expression in hearts of transgenic mice. Mice expressing beta2 had a severe phenotype with juvenile lethality. Myofibril architecture was severely disrupted. The beta2 did not localize to the Z-line. Therefore, beta1 has a distinct function that includes interactions at the Z-line. Mice expressing beta1 showed altered morphology of the intercalated disc, without the lethality or myofibril disruption of the beta2-expressing mice. The in vivo function of CP is presumed to involve binding barbed ends of actin filaments. To test this hypothesis, we expressed a beta1 mutant that poorly binds actin. These mice showed both myofibril disruption and intercalated disc remodeling, as predicted. Therefore, CPbeta1 and CPbeta2 each have a distinct function that cannot be provided by the other isoform. CPbeta1 attaches actin filaments to the Z-line, and CPbeta2 organizes the actin at the intercalated discs.

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The expression of CPβ1, CPβ1-L262R, CPβ2, and CPα in TG-β1L262R lines. The expression of TG-β1L262R lines was determined by two-dimensional immunoblots probed with anti-panα, anti-β1, and anti-β2. The β1 and β1-L262R polypeptides have a similar molecular mass, but different pI's, 5.5 and 5.8, respectively.
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Figure 4: The expression of CPβ1, CPβ1-L262R, CPβ2, and CPα in TG-β1L262R lines. The expression of TG-β1L262R lines was determined by two-dimensional immunoblots probed with anti-panα, anti-β1, and anti-β2. The β1 and β1-L262R polypeptides have a similar molecular mass, but different pI's, 5.5 and 5.8, respectively.

Mentions: To determine the CPβ isoform protein expression in TG-β1L262R lines, two-dimensional immunoblots were required to identify the β isoforms (Fig. 4). Although the β1 and β1-L262R polypeptides have similar molecular masses, the β1-L262R mutation shifts the pI from 5.5 to 5.8 and then can be discriminated from wild-type (see Fig. 4, cartoon). The blots were also probed with an mAb pan-reactive to α1 and α2.


Vertebrate isoforms of actin capping protein beta have distinct functions In vivo.

Hart MC, Cooper JA - J. Cell Biol. (1999)

The expression of CPβ1, CPβ1-L262R, CPβ2, and CPα in TG-β1L262R lines. The expression of TG-β1L262R lines was determined by two-dimensional immunoblots probed with anti-panα, anti-β1, and anti-β2. The β1 and β1-L262R polypeptides have a similar molecular mass, but different pI's, 5.5 and 5.8, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2168092&req=5

Figure 4: The expression of CPβ1, CPβ1-L262R, CPβ2, and CPα in TG-β1L262R lines. The expression of TG-β1L262R lines was determined by two-dimensional immunoblots probed with anti-panα, anti-β1, and anti-β2. The β1 and β1-L262R polypeptides have a similar molecular mass, but different pI's, 5.5 and 5.8, respectively.
Mentions: To determine the CPβ isoform protein expression in TG-β1L262R lines, two-dimensional immunoblots were required to identify the β isoforms (Fig. 4). Although the β1 and β1-L262R polypeptides have similar molecular masses, the β1-L262R mutation shifts the pI from 5.5 to 5.8 and then can be discriminated from wild-type (see Fig. 4, cartoon). The blots were also probed with an mAb pan-reactive to α1 and α2.

Bottom Line: The beta2 did not localize to the Z-line.Therefore, CPbeta1 and CPbeta2 each have a distinct function that cannot be provided by the other isoform.CPbeta1 attaches actin filaments to the Z-line, and CPbeta2 organizes the actin at the intercalated discs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Actin capping protein (CP) binds barbed ends of actin filaments to regulate actin assembly. CP is an alpha/beta heterodimer. Vertebrates have conserved isoforms of each subunit. Muscle cells contain two beta isoforms. beta1 is at the Z-line; beta2 is at the intercalated disc and cell periphery in general. To investigate the functions of the isoforms, we replaced one isoform with another using expression in hearts of transgenic mice. Mice expressing beta2 had a severe phenotype with juvenile lethality. Myofibril architecture was severely disrupted. The beta2 did not localize to the Z-line. Therefore, beta1 has a distinct function that includes interactions at the Z-line. Mice expressing beta1 showed altered morphology of the intercalated disc, without the lethality or myofibril disruption of the beta2-expressing mice. The in vivo function of CP is presumed to involve binding barbed ends of actin filaments. To test this hypothesis, we expressed a beta1 mutant that poorly binds actin. These mice showed both myofibril disruption and intercalated disc remodeling, as predicted. Therefore, CPbeta1 and CPbeta2 each have a distinct function that cannot be provided by the other isoform. CPbeta1 attaches actin filaments to the Z-line, and CPbeta2 organizes the actin at the intercalated discs.

Show MeSH
Related in: MedlinePlus