CKIα Is Associated with and Phosphorylates Star-PAP and Is Also Required for Expression of Select Star-PAP Target Messenger RNAs

Abstract

We have recently identified Star-PAP, a nuclear poly(A) polymerase that associates with phosphatidylinositol-4-phosphate 5-kinase I␣ (PIPKI␣) and is required for the expression of a specific subset of mRNAs. Star-PAP activity is directly modulated by the PIPKI␣ product phosphatidylinositol 4,5-bisphosphate (PI-4,5-P 2 ), linking nuclear phosphoinositide signaling to gene expression. Here, we show that PI-4,5-P 2 -dependent protein kinase activity is also a part of the Star-PAP protein complex. We identify the PI-4,5-P 2 -sensitive casein kinase I␣ (CKI␣) as a protein kinase responsible for this activity and further show that CKI␣ is capable of directly phosphorylating Star-PAP. Both CKI␣ and PIPKI␣ are required for the synthesis of some but not all Star-PAP target mRNA, and like Star-PAP, CKI␣ is associated with these messages in vivo. Taken together, these data indicate that CKI␣, PIPKI␣, and Star-PAP function together to modulate the production of specific Star-PAP messages. The Star-PAP complex therefore represents a location where multiple signaling pathways converge to regulate the expression of specific mRNAs.Polyadenylation of most mRNAs is required for their efficient transcription and export as well as regulating their stability and translational efficiency (1). Polyadenylation of mRNA is achieved through the activity of poly(A) polymerases (PAPs). 2There are multiple PAPs in mammalian cells, including PAP␣, which is thought to be primarily responsible for the polyadenylation of newly transcribed mRNAs in the nucleus (2, 3). Additionally, there are "non-canonical" PAPs, including Gld2 and Trf4, which regulate stability and degradation of specific RNAs through polyadenylation (4, 5).Star-PAP is a non-canonical poly(A) polymerase that is distinct from all other currently characterized PAP enzymes (6). Like the canonical PAP␣, Star-PAP has an RNA recognition motif and is a nuclear enzyme. However, similar to non-canonical PAPs, the Star-PAP protein complex and architecture differ significantly from PAP␣, and consequently, Star-PAP specifically targets a select subset of mRNAs. This suggests that Star-PAP is a hybrid PAP that is required for the 3Ј-end formation of newly transcribed pre-mRNAs but functions in a regulatory role to control mRNA expression levels. Star-PAP has a unique domain structure relative to all other known PAPs (6). One unique feature is a 205-amino acid proline-rich region (PRR) inserted into the catalytic PAP core. The PRR splits the catalytic PAP domain, and this region represents a potential site for regulation of Star-PAP function.Poly(A) polymerases operate as large multiprotein complexes responsible for the 3Ј-processing of RNAs (7, 8). The Star-PAP polyadenylation complex is similar to that of poly(A) polymerases that utilizes mRNA as a substrate and includes cleavage and polyadenylation specificity factor (CPSF) subunits, cleavage stimulatory factor (CstF) subunits, symplekin, and RNA polymerase II (6, 9). However, unlike other poly(A) polymerase complexes, the Star-PAP complex...

Effect of heme oxygenase-1 on vascular function and disease

“…The authors suggested that antioxidant response induces the assembly of the Star-PAP complex, leading to a rapid initiation of 3 0 -end formation and polyadenylation by the Star-PAP complex. Moreover, the PI4,5P 2 sensitive kinase CKIa is important for the production of the Star-PAP target mRNAs, including HO-1, however, in contrast to PIPKIa, CKIa is not necessary for HO-1 induction but only for basal expression [5]. Whether there is any link between Star-PAP complex and Nrf2 transcription factor is not known yet, although the involvement of STAR-PAP in tBHQ-induced expression of HO-1 suggests such an interaction, as tBHQ is a potent activator of Nrf2.…”

Inhibition of polyadenylation reduces inflammatory gene induction

“…However, in a proteomic study of the mRNA polyadenylation complex, only its close relative PAPg was recovered (Shi et al 2009). In addition, it has recently transpired that a subset of mRNAs are polyadenylated by the more distantly related poly(A) polymerase called Star-PAP/PAPD2 in response to specific signal transduction (Gonzales et al 2008). Strikingly, 80% knockdown of PAPa only has a small effect on the poly(A) population in the cell, but still causes a significant reduction in the induction of CXCL1, CCL2, and IL8 mRNA, suggesting that PAPa is perhaps more important for the expression of inflammatory genes than for general gene expression (Fig.…”

PIP kinases define PI4,5P2 signaling specificity by association with effectors

“…Moreover, nuclear PI4,5P 2 also functions as a direct messenger that binds and regulates nuclear PI4,5P 2 effectors. Recent progress has identified a set of nuclear PI4,5P 2 effectors directly involved in multiple processes, including transcription [177–180], mRNA processing [181–185], mRNA export [186, 187], chromatin remodeling [180], stress responses [185], DNA repair, and mitosis [188]. The PIP kinases responsible for PI4,5P 2 generation in the nuclear envelope are not clear, but the intra-nuclear PIP kinases and PI4,5P 2 signaling are more defined.…”

“…The PI4,5P 2 generated by PIPKIα appears to directly activate Star-PAP and also regulates other components of the Star-PAP complex, as exemplified by the Ser/Thr kinase casein kinase I (CKI) and protein kinase C δ (PKC5), both of which are required to activate Star-PAP for mRNA processing [181, 182, 185]. The functionally redundant CKI isoforms α and ε phosphorylate Star-PAP at the proline rich region (PRR) upon oxidative stress, which is critical for Star-PAP polyadenylation activity.…”