Nonmyristoylated MARCKS Complements Some but Not All of the Developmental Defects Associated with MARCKS Deficiency in Mice

Abstract

The myristoylated alanine-rich C kinase substrate, or MARCKS protein, is a widely expressed, prominent substrate for protein kinase C. Although the exact function of MARCKS has not been elucidated, targeted disruption of the MARCKS gene (Macs) in mice has shown that MARCKS plays a crucial role in the development of the central nervous system. Mice deficient in MARCKS exhibited universal perinatal death with defects in neurulation, fusion of the cerebral hemispheres, formation of the great forebrain commissures, and retinal and cortical lamination (Stumpo et al., Proc. Natl. Acad. Sci. USA 92, 944-948, 1995). In the present studies, a transgene consisting of approximately 3.4 kb of promoter from the human MARCKS gene (MACS), with an epitope tag sequence inserted at the carboxyl terminus of the MARCKS coding region, was able to complement completely MARCKS deficiency in mice. Thus, the human transgene contained all of the elements necessary for normal developmental expression of MARCKS. To test the importance of MARCKS myristoylation to its developmental role, an otherwise identical transgene was constructed in which the glycine at the amino terminus of MARCKS was mutated to an alanine. This mutation, which resulted in the expression of nonmyristoylated MARCKS, was successful in partially rescuing the Macs null phenotype. Specifically, about 25% of these mice survived the perinatal period; these survivors appeared to develop normally except for slightly decreased body size. In both the survivors and the nonsurvivors, all of the known anatomical defects associated with MARCKS deficiency were corrected by expression of the nonmyristoylated human protein. These results indicate that myristoylation of MARCKS is not required for the protein to correct many of the developmental abnormalities characteristic of its deficiency.

Functional Role of the Interaction between Polysialic Acid and Myristoylated Alanine-rich C Kinase Substrate at the Plasma Membrane

“…6A), whereas labeled colominic acid/PSA did not bind to cells expressing GFP fusions with the non-myristoylatable MARCKS mutant (Fig. 6B) or the nonphosphorylatable MARCKS mutants (data not shown), neither of which insert into membranes (19,28). Similarly, colominic acid/PSA labeled with the dye HiLyte Fluor 405 also bound to cells expressing non-mutated MARCKS-GFP (Fig.…”

“…MARCKS is a protein kinase C (PKC)-specific substrate and binds calmodulin and actin (15,16). MARCKS is a rod-shaped, acidic protein that possesses three highly conserved regions: 1) the N terminus, which contains a consensus sequence for myristoylation that is involved in membrane binding of MARCKS (17), 2) the MH2 domain, which resembles the cytoplasmic tail of the cation-independent mannose-6-phosphate receptor, and 3) the phosphorylation site domain, which contains the PKC phosphorylation sites and the effector domain (18,19). The effector domain is highly basic, in contrast to the rest of the highly acidic protein, and has been shown to be crucial for the function of MARCKS, whereas myristoylation of MARCKS is not required for many of the in vivo functions of MARCKS as indicated by studies using expression of non-myristoylatable MARCKS in MARCKS-null mice (19).…”

Myristoylated Alanine‐Rich Protein Kinase Substrate (MARCKS) Regulates Small GTPase Rac1 and Cdc42 Activity and Is a Critical Mediator of Vascular Smooth Muscle Cell Migration in Intimal Hyperplasia Formation

“…A schematic representation of WT and MARCKS mutants is shown in Figure 4A. These plasmids and the function of MARCKS mutants have been well characterized 13, 36, 37, 38. The G2A mutant lacks the myristoylated tail, the S4G mutant expresses a phosphorylation‐deficient ED domain, and the S4D has a pseudophosphorylated ED domain.…”

Molecular basis of lithium action: integration of lithium-responsive signaling and gene expression networks

“…87 MARCKS is preferentially expressed in dendritic branches and axon terminals within limbic and limbic-associated regions of brain, as well as neuronal growth cones necessary for normal brain development. [88][89][90] Within brain and neurons in culture, MARCKS protein is expressed in neurites and synaptosomes, and is colocalized with synaptic vesicles. [91][92][93] PKC-MARCKS signaling has been implicated in vesicular trafficking of neurotransmitter in living neurons, 94 and in the regulation of various processes including synaptic transmission at nerve terminals.…”