RNA Binding Protein RBM38 Regulates Expression of the 11-Kilodalton Protein of Parvovirus B19, Which Facilitates Viral DNA Replication

Abstract

Human parvovirus B19 (B19V) expresses a single precursor mRNA (pre-mRNA), which undergoes alternative splicing and alternative polyadenylation to generate 12 viral mRNA transcripts that encode two structural proteins (VP1 and VP2) and three nonstructural proteins (NS1, 7.5-kDa protein, and 11-kDa protein). Splicing at the second 5' donor site (D2 site) of the B19V pre-mRNA is essential for the expression of VP2 and the 11-kDa protein. We previously identified that -acting intronic splicing enhancer 2 (ISE2) that lies immediately after the D2 site facilitates the recognition of the D2 donor for its efficient splicing. In this study, we report that ISE2 is critical for the expression of the 11-kDa viral nonstructural protein. We found that ISE2 harbors a consensus RNA binding motif protein 38 (RBM38) binding sequence, 5'-UGUGUG-3'. RBM38 is expressed during the middle stage of erythropoiesis. We first confirmed that RBM38 binds specifically with the ISE2 element The knockdown of RBM38 significantly decreases the level of spliced mRNA at D2 that encodes the 11-kDa protein but not that of the D2-spliced mRNA that encodes VP2. Importantly, we found that the 11-kDa protein enhances viral DNA replication and virion release. Accordingly, the knockdown of RBM38 decreases virus replication via downregulating 11-kDa protein expression. Taken together, these results suggest that the 11-kDa protein facilitates B19V DNA replication and that RBM38 is an essential host factor for B19V pre-mRNA splicing and for the expression of the 11-kDa protein. B19V is a human pathogen that can cause fifth disease, arthropathy, anemia in immunocompromised patients and sickle cell disease patients, myocarditis, and hydrops fetalis in pregnant women. Human erythroid progenitor cells (EPCs) are most susceptible to B19V infection and fully support viral DNA replication. The exclusive tropism of B19V for erythroid-lineage cells is dependent not only on the expression of viral receptors and coreceptors on the cell surface but also on the intracellular host factors that support B19V replication. Our present study shows that B19V uses a host factor, RNA binding motif protein 38 (RBM38), for the processing of its pre-mRNA during virus replication. Specifically, RBM38 interacts with the intronic splicing enhancer 2 (ISE2) element of B19V pre-mRNA and promotes 11-kDa protein expression, thereby regulating the 11-kDa protein-mediated augmentation of B19V replication. The identification of this novel host-pathogen interaction will provide mechanistic insights into B19V replication and aid in finding new targets for anti-B19V therapeutics.

RNA Binding Motif Protein RBM45 Regulates Expression of the 11-Kilodalton Protein of Parvovirus B19 through Binding to Novel Intron Splicing Enhancers

“…Identification of B19V ISE2-interacting proteins. We previously reported that ISE2 is critical for the recognition of the D2 site for splicing and that the interaction of RBM38 with ISE2 promotes splicing of the intron from the D2 to the A2-2 splice sites (37,41). Thus, we asked whether other host factors could interact with ISE2 and facilitate the alternative splicing at these sites.…”

“…The viral precursor mRNA (pre-mRNA) is alternatively spliced and alternatively polyadenylated resulting in the generation of nine major viral mRNA transcripts that encode capsid proteins (VP1 and VP2) and nonstructural proteins (NS1, 11-kDa, and 7.5-kDa) (31)(32)(33)(34). NS1 is the only viral protein that is essential for B19V DNA replication (35)(36)(37). However, the small nonstructural protein 11-kDa, which is localized predominantly in the cytoplasm and plays an enhancement role in viral DNA replication, is unique among parvoviruses (38).…”

“…The intronic splicing enhancer 2 (ISE2), located immediately downstream of the second donor (D2), is critical for splicing of the viral pre-mRNA at D2, which controls production of the mRNAs encoding both VP2 and 11-kDa (41). We have shown that RNA binding motif protein 38 (RBM38) binds to ISE2 and executes a function in splicing of viral pre-mRNA at D2 and the second acceptor of the second intron (A2-2), which encodes mRNA encoding 11-kDa (37). However, how the RBM38 specifically regulates maturation of the 11-kDa-encoding mRNA is unknown.…”

Genetic Basis of Myocarditis: Myth or Reality?

“…It may also infect non-erythroid lineage cells such as myocardial endothelial cells [60]. The tropism of B19V for EPCs depends on the expression of viral receptors and coreceptors on the cell surface and by the viral use of host factors such as RNA binding motif protein 38 (RBM38) for the processing of its pre-mRNA during viral replication [61]. Variants in RBM38 gene have not been reported so far.…”

New Insights of Human Parvovirus B19 in Modulating Erythroid Progenitor Cells Differentiation

“…The region of them shows a greater sequence variation in contrast to the large nonstructural protein(NS1) region that is highly conserved, which implies their function in host antivirus responses [28]. Expressions of the other two small nonstructural proteins of 11 kDa and 7.5 kDa were also documented [29,30], the former one was suggested as a potent inducer of apoptosis via enhancing viral DNA replication and virion release [31,32], while the function of 7.5 kDa protein is inconclusive.…”