Inactive forms of the catalytic subunit of protein kinase A are expressed in the brain of higher primates

Abstract

It is well documented that the β‐gene of the catalytic (C) subunit of protein kinase A encodes a number of splice variants. These splice variants are equipped with a variable N‐terminal end encoded by alternative use of several exons located 5′ to exon 2 in the human, bovine and mouse Cβ gene. In the present study, we demonstrate the expression of six novel human Cβ mRNAs that lack 99 bp due to loss of exon 4. The novel splice variants, designated CβΔ4, were identified in low amounts at the mRNA level in NTera2‐N cells. We developed a method to detect CβΔ4 mRNAs in various cells and demonstrated that these variants were expressed in human and Rhesus monkey brain. Transient expression and characterization of the CβΔ4 variants demonstrated that they are catalytically inactive both in vitro against typical protein kinase A substrates such as kemptide and histone, and in vivo against the cAMP‐responsive element binding protein. Furthermore, co‐expression of CβΔ4 with the regulatory subunit (R) followed by kinase activity assay with increasing concentrations of cAMP and immunoprecipitation with extensive washes with cAMP (1 mm) and immunoblotting demonstrated that the CβΔ4 variants associate with both RI and RII in a cAMP‐independent fashion. Expression of inactive C subunits which associate irreversibly with R may imply that CβΔ4 can modulate local cAMP effects in the brain by permanent association with R subunits even at saturating concentrations of cAMP.

Protein kinase A type I activates a CRE-element more efficiently than protein kinase A type II regardless of C subunit isoform

“…PKA-specific catalytic activity was determined as described previously [66]. Molar amounts of C subunit were determined assuming 600 pmol phosphate transferred per min per mg pure bovine C.…”

Evolutionary Paths of the cAMP-Dependent Protein Kinase (PKA) Catalytic Subunits

“…In addition, Cβ variants formed by skipping of exon 4 are expressed in the brain of higher primates. These C subunits bind the R subunit in a cAMP-independent fashion [34].…”

Evolution of the cAMP-dependent protein kinase (PKA) catalytic subunit isoforms

“…These sequence differences may be associated with functional differences and hence in part explain why they are both universally conserved in the bony vertebrates [22]. With the exception of brain-specific deletion mutants in PRKACB lacking exon 4 (CβΔ4) [23], all Cα and Cβ proteins share the same conserved core in the segment known as Core 16-350 , consisting of Cα1/Cβ1 residues 16 to 350. These residues are encoded by PRKACA and PRKACB paralogous exons 2 to 10 [22].…”

“…Exons 1–3 and 1–4 encode the neural-specific proteins Cβ3 and Cβ4, respectively. Exons a, b, and c are only seen in combination with exons 1–3 or 1–4, and are therefore also neural-specifically expressed [8, 23, 39, 40, 43]. …”