Identification of a calcium-dependent calmodulin-binding domain in Xenopus membrane skeleton protein 4.1

Membrane skeleton protein 4.1 and other members of a family of proteins that link the cytoskeleton to the plasma membrane may play an integral role in cell communication during development. The polymerase chain reaction and degenerate oligodeoxynucleotide primers to consensus sequences in the putative membrane-binding domain of the protein 4.1 superfamily were used to isolate cDNAs encoding members of the zebrafish protein 4.1 family. Zebrafish stage- and tissue-specific first strand cDNA was used in the PCR. After the reaction, amplicons of the predicted size were sequenced to confirm their relationship to the protein 4.1 superfamily. One cDNA, with a high degree of similarity to a mouse novel band 4.1-like cDNA, was used to probe a zebrafish adult brain library. A 2.4-kb cDNA was isolated and found to encode a 619 amino acid polypeptide homologous to mouse novel band 4.1-like protein 4. Zebrafish nbl4 mRNA is maternally supplied and is expressed throughout embryogenesis. In adults, nbl4 is found in the ovary, eye, heart, and brain, but not in gut or skeletal muscle. When synthetic nbl4 mRNA is translated in vitro it binds calmodulin in a calcium-dependent manner. These data indicate that zebrafish nbl4 is a maternal transcript owing to its presence before the midblastula transition, and it is present later on in specific adult structures. The ability to bind calmodulin would suggest that the function of nbl4 protein may be potentially regulated via a calcium-calmodulin dependent mechanism.

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Phosphorylation of serine residues affects the conformation of the calmodulin binding domain of human protein 4.1

European Journal of Biochemistry ◽

10.1046/j.1432-1327.2001.02347.x ◽

2001 ◽

Vol 268 (15) ◽

pp. 4292-4299 ◽

Cited By ~ 10

Author(s):

Stefan W. Vetter ◽

Estelle Leclerc

Keyword(s):

Binding Domain ◽

Human Protein ◽

Protein 4.1 ◽

Calmodulin Binding

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Calcium-Dependent and -Independent Interactions of the Calmodulin-Binding Domain of Cyclic Nucleotide Phosphodiesterase with Calmodulin†

Biochemistry ◽

10.1021/bi9816453 ◽

1999 ◽

Vol 38 (5) ◽

pp. 1446-1455 ◽

Cited By ~ 46

Author(s):

Tao Yuan ◽

Michael P. Walsh ◽

Cindy Sutherland ◽

Heinz Fabian ◽

Hans J. Vogel

Keyword(s):

Cyclic Nucleotide ◽

Binding Domain ◽

Cyclic Nucleotide Phosphodiesterase ◽

Calcium Dependent ◽

Calmodulin Binding

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The carboxyterminal EF domain of erythroid α-spectrin is necessary for optimal spectrin-actin binding

Blood ◽

10.1182/blood-2009-12-260612 ◽

2010 ◽

Vol 116 (14) ◽

pp. 2600-2607 ◽

Cited By ~ 11

Author(s):

Catherine Korsgren ◽

Samuel E. Lux

Keyword(s):

Membrane Skeleton ◽

Binding Domain ◽

Actin Binding ◽

Binary Interaction ◽

Spectrin Repeat ◽

Calcium Dependent ◽

Actin Binding Domain ◽

Ef Hands ◽

Protein 4.1R ◽

Skeletal Integrity

Abstract Spectrin and protein 4.1R crosslink F-actin, forming the membrane skeleton. Actin and 4.1R bind to one end of β-spectrin. The adjacent end of α-spectrin, called the EF domain, is calmodulin-like, with calcium-dependent and calcium-independent EF hands. The severely anemic sph1J/sph1J mouse has very fragile red cells and lacks the last 13 amino acids in the EF domain, implying that the domain is critical for skeletal integrity. To test this, we constructed a minispectrin heterodimer from the actin-binding domain, the EF domain, and 4 adjacent spectrin repeats in each chain. The minispectrin bound to F-actin in the presence of native human protein 4.1R. Formation of the spectrin-actin-4.1R complex was markedly attenuated when the minispectrin contained the shortened sph1J α-spectrin. The α-spectrin deletion did not interfere with spectrin heterodimer assembly or 4.1R binding but abolished the binary interaction between spectrin and F-actin. The data show that the α-spectrin EF domain greatly amplifies the function of the β-spectrin actin-binding domain (ABD) in forming the spectrin-actin-4.1R complex. A model, based on the structure of α-actinin, suggests that the EF domain modulates the function of the ABD and that the C-terminal EF hands (EF34) may bind to the linker that connects the ABD to the first spectrin repeat.

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Calcium-dependent and -independent Binding of Soybean Calmodulin Isoforms to the Calmodulin Binding Domain of Tobacco MAPK Phosphatase-1

Journal of Biological Chemistry ◽

10.1074/jbc.m608970200 ◽

2007 ◽

Vol 282 (9) ◽

pp. 6031-6042 ◽

Cited By ~ 25

Author(s):

Mario Rainaldi ◽

Aaron P. Yamniuk ◽

Tomohiko Murase ◽

Hans J. Vogel

Keyword(s):

Binding Domain ◽

Calcium Dependent ◽

Calmodulin Binding ◽

Mapk Phosphatase

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Similarities and differences in the structure and function of 4.1G and 4.1R135, two protein 4.1 paralogues expressed in erythroid cells

Biochemical Journal ◽

10.1042/bj20100041 ◽

2010 ◽

Vol 432 (2) ◽

pp. 407-416 ◽

Cited By ~ 7

Author(s):

Wataru Nunomura ◽

Kengo Kinoshita ◽

Marilyn Parra ◽

Philippe Gascard ◽

Xiuli An ◽

...

Keyword(s):

Membrane Proteins ◽

Erythroid Differentiation ◽

Band 3 ◽

Erythroid Cells ◽

Protein 4.1 ◽

Calcium Dependent ◽

Calmodulin Binding ◽

And Function ◽

Glycophorin C ◽

In Erythroid Cells

Membrane skeletal protein 4.1R is the prototypical member of a family of four highly paralogous proteins that include 4.1G, 4.1N and 4.1B. Two isoforms of 4.1R (4.1R135 and 4.1R80), as well as 4.1G, are expressed in erythroblasts during terminal differentiation, but only 4.1R80 is present in mature erythrocytes. Although the function of 4.1R isoforms in erythroid cells has been well characterized, there is little or no information on the function of 4.1G in these cells. In the present study, we performed detailed characterization of the interaction of 4.1G with various erythroid membrane proteins and the regulation of these interactions by calcium-saturated calmodulin. Like both isoforms of 4.1R, 4.1G bound to band 3, glycophorin C, CD44, p55 and calmodulin. While both 4.1G and 4.1R135 interact with similar affinity with CD44 and p55, there are significant differences in the affinity of their interaction with band 3 and glycophorin C. This difference in affinity is related to the non-conserved N-terminal headpiece region of the two proteins that is upstream of the 30 kDa membrane-binding domain that harbours the binding sites for the various membrane proteins. The headpiece region of 4.1G also contains a high-affinity calcium-dependent calmodulin-binding site that plays a key role in modulating its interaction with various membrane proteins. We suggest that expression of the two paralogues of protein 4.1 with different affinities for band 3 and glycophorin C is likely to play a role in assembly of these two membrane proteins during terminal erythroid differentiation.

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