Emerin Is Hyperphosphorylated and Redistributed in Herpes Simplex Virus Type 1-Infected Cells in a Manner Dependent on both UL34 and US3

Abstract

Cells infected with wild-type herpes simplex virus type 1 (HSV-1) show disruption of the organization of the nuclear lamina that underlies the nuclear envelope. This disruption is reflected in changes in the localization and phosphorylation of lamin proteins. Here, we show that HSV-1 infection causes relocalization of the LEM domain protein emerin. In cells infected with wild-type virus, emerin becomes more mobile in the nuclear membrane, and in cells infected with viruses that fail to express UL34 protein (pUL34) and US3 protein (pUS3), emerin no longer colocalizes with lamins, suggesting that infection causes a loss of connection between emerin and the lamina. Infection causes hyperphosphorylation of emerin in a manner dependent upon both pUL34 and pUS3. Some emerin hyperphosphorylation can be inhibited by the protein kinase C␦ (PKC␦) inhibitor rottlerin. Emerin and pUL34 interact physically, as shown by pull-down and coimmunoprecipitation assays. Emerin expression is not, however, necessary for infection, since virus growth is not impaired in cells derived from emerin-null transgenic mice. The results suggest a model in which pUS3 and PKC␦ that has been recruited by pUL34 hyperphosphorylate emerin, leading to disruption of its connections with lamin proteins and contributing to the disruption of the nuclear lamina. Changes in emerin localization, nuclear shape, and lamin organization characteristic of cells infected with wild-type HSV-1 also occur in cells infected with recombinant virus that does not make viral capsids, suggesting that these changes occur independently of capsid envelopment.During primary envelopment, herpes simplex virus type 1 (HSV-1) nucleocapsids translocate from the nucleus to the cytoplasm by budding into the inner nuclear membrane and then fusing with the outer nuclear membrane. The capsid does not, however, have unimpeded access to the inner nuclear membrane. Lining the inside of the inner nuclear membrane is the nuclear lamina, which is composed of a meshwork of proteins with spaces too small for the capsid to move through without some disruption (2,19,65). The lamina meshwork is made up of intermediate filament family proteins called lamins that are linked to the inner nuclear membrane and to intranuclear proteins by association with lamin-associated proteins (LAPs) (reviewed in reference 65). Connection of the network of lamin proteins to the inner nuclear membrane is mediated by integral membrane LAPs, including emerin, lamin B receptor, LAP2-␤, and MAN-1 (26).Emerin is a member of a family of nuclear envelope proteins that share a common sequence called the LEM domain that mediates association with BAF (barrier to autointegration factor) and is important for the assembly of LEM domain proteins into the re-forming nuclear envelope following mitosis (21,37,39). Emerin also contains a lamin-binding domain that helps retain it in the interphase nuclear envelope (6,14,25,37). Emerin is ubiquitously expressed but is not essential for the viability of cells in culture (36). Failure to ...

Cytomegaloviral proteins that associate with the nuclear lamina: components of a postulated nuclear egress complex

“…PKC and pUL97. There is accumulating evidence that the involvement of both cellular and viral protein kinases (some of which possess direct lamin-phosphorylating activity) is important for the regulation of nuclear egress of other herpesviruses as well (Klupp et al, 2001;Reynolds et al, 2002;Marschall et al, 2005;Bjerke & Roller, 2006;Park & Baines, 2006;Kato et al, 2006;Leach et al, 2007). It will be crucial to determine the exact phosphorylation target sites on lamins and lamina-associated proteins of these protein kinases for a detailed understanding of the cytomegaloviral nuclear egress.…”

“…Recruitment of PKC to the nuclear membrane occurs during infection with HCMV, murine cytomegalovirus (MCMV) and herpes simplex virus type-1 (HSV-1) (Muranyi et al, 2002;Park & Baines, 2006;Milbradt et al, 2007). A conserved group of herpesvirus-encoded lamina-associated proteins have been implicated in the recruitment of PKC, the rearrangement of nuclear lamina components and viral nuclear egress (Reynolds et al, 2004;Muranyi et al, 2002;Leach et al, 2007;Morris et al, 2007;Fuchs et al, 2002;Farina et al, 2005;Milbradt et al, 2007; Camozzi et al, 2008). This group comprises pUL50 and pUL53 in HCMV and their homologues in other herpesviruses.…”

Role of tegument proteins in herpesvirus assembly and egress

“…Alphaherpesviruses also express a viral serine/threonine protein kinase, pUS3, that phosphorylates lamins and lamin receptor emerin, which are thought to be involved in maintaining nuclear integrity (Leach et al, 2007;Morris et al, 2007;Mou et al, 2008). In addition, both HSV-1 and PrV pUL31 and pUL34 have been shown to be phosphorylated in a pUS3-dependent manner, and all three proteins have been found to be incorporated into primary virions (Purves et al, 1991; Granzow et al, 2004;Ryckman and Roller, 2004;Kato et al, 2005;Mou et al, 2007Mou et al, , 2009.…”

“…HSV-1 and PrV disrupt the lamina through two viral proteins, pUL31 and pUL34, which form a complex colocalized to the inner NM (Reynolds et al, 2002;Reynolds et al, 2004;Simpson-Holley et al, 2004;Simpson-Holley et al, 2005;Bjerke and Roller, 2006). Research suggested that pUL31 and pUL34 promote viral nuclear egress by several mechanisms, including (1) affecting maturation of viral replication intermediates so that capsids assemble adjacent to the nuclear envelope, making it convenient for primary egress (Simpson-Holley et al, 2004, (2) causing displacement of and conformational changes in lamins A/C and B (Reynolds et al, 2004;Simpson-Holley et al, 2004;Bjerke and Roller, 2006), and (3) mislocalizing the membrane lamin receptors, such as the lamin B receptor and emerin, which tether lamins to the inner NM (Scott and O'Hare, 2001;Leach et al, 2007;Morris et al, 2007;Mou et al, 2008). Amazingly, expression of PrV pUL31 and pUL34 in RK-13 cells resulted in formation of vesicles in the perinuclear space, indicating that they may compose the essential budding machinery (Klupp et al, 2007).…”

“…There, the viruses encounter the nuclear lamina, a rigid network of lamin proteins lining the inner NM, which is a major obstacle for budding of the nucleocapsids. Herpesviruses disrupt the nuclear lamina in order to bud into the perinuclear space and acquire the inner NM as the envelope, the so called primary envelopment (Scott and O'Hare, 2001;Muranyi et al, 2002;Leach et al, 2007;Mou et al, 2007;Mou et al, 2008). HSV-1 and PrV disrupt the lamina through two viral proteins, pUL31 and pUL34, which form a complex colocalized to the inner NM (Reynolds et al, 2002;Reynolds et al, 2004;Simpson-Holley et al, 2004;Simpson-Holley et al, 2005;Bjerke and Roller, 2006).…”