Analysis of akap12 in zebrafish: spatiotemporal expression of two isoforms and pancreas development

Abstract

A Kinase Anchoring Protein 12 (AKAP12), also known as Gravin and src-suppressed C kinase substrate (SSeCKS), is a multivalent scaffolding protein that controls multiple biological processes. Its activity results from scaffolding signaling proteins such as protein kinase (PK) C and A, calmodulin, cyclins, phosphoinositides, β-1,4 galactosyltransferase, Src, as well as the actin cytoskeleton. It has been demonstrated that there are three types of AKAP12 isoforms (designated α, β, and γ) in human with different promoters and known that the major types are AKAP12α and AKAP12β. In zebrafish, the expression of AKAP12 is dynamically controlled. It is detected ubiquitously during gastrulation. During segmentation, the expression is restricted to the mesoderm, nervous system and blood vasculature. Later, it becomes localized to the head region. The composition and distribution of AKAP12 indicates its crucial role and function in development process. However, similarity and difference of expression and distribution between AKAP12α and AKAP12β in zebrafish remain unclear. Here we investigated the biological significance of these two isoforms during zebrafish development. During zebrafish embryogenesis, RT-PCR analysis detected the mRNA expression of AKAP12 isoforms at different developmental stages: AKAP12β is detected from the late blastula period

Expression of AKAP12α is initiated at bud stage, the late gastrula period. By knockdown of each isoform expression using morpholino-based antisense oligonucleotide, respective expression of AKAP12α and AKAP12β was identified. To clarify the pattern of expression clearly, we designed probes for each isoforms. As a result, zebrafish AKAP12α, β were all found to be expressed similarly but distinctly. Taken together, these observations become essential for identifying the role and function of AKAP12α and AKAP12β that are involved in the zebrafish development.

In addition to identify spatiotemporal expression of the two isoforms, we uncovered the role of akap12 in pancreas laterality. During pancreas development, the organ primordia undergo asymmetric movement to ensure proper function. Yet the role of akap12 in pancreas laterality is not well defined. In this study, we investigated the function of akap12 in asymmetric movement of pancreas during development. In the absence of akap12, we identified disrupted pancreas morphology. In particular, akap12 does not function directly in pancreas development. We confirmed the result by KV-specific knock-down technic. Collectively, our data provide the role of akap12 in pancreas laterality.