Velcro in the nuclear envelope: LBR and LAPs

Human papillomavirus (HPV) infections of the squamous epithelium are associated with high-level expression of the E1 E4 protein during the productive phase of infection. However, the precise mechanisms of how E1 E4 contributes to the replication cycle of the virus are poorly understood. Here, we show that the serine-arginine (SR)-specific protein kinase SRPK1 is a novel binding partner of HPV type 1 (HPV1) E1 E4. We map critical residues within an arginine-rich domain of HPV1 E1 E4, and in a region known to facilitate E1 E4 oligomerization, that are requisite for SRPK1 binding. In vitro kinase assays show that SRPK1 binding is associated with phosphorylation of an HPV1 E1 E4 polypeptide and modulates autophosphorylation of the kinase. We show that SRPK1 is sequestered into E4 inclusion bodies in terminally differentiated cells within HPV1 warts and that colocalization between E1 E4 and SRPK1 is not dependent on additional HPV1 factors. Moreover, we also identify SRPK1 binding of E1 E4 proteins of HPV16 and HPV18. Our findings indicate that SRPK1 binding is a conserved function of E1 E4 proteins of diverse virus types. SRPK1 influences important biochemical processes within the cell, including nuclear organization and RNA metabolism. While phosphorylation of HPV1 E4 by SRPK1 may directly influence HPV1 E4 function during the infectious cycle, the modulation and sequestration of SRPK1 by E1 E4 may affect the ability of SRPK1 to phosphorylate its cellular targets, thereby facilitating the productive phase of the HPV replication cycle.Human papillomaviruses (HPVs) comprise a family of small double-stranded DNA viruses that have a tropism for human epithelial cells at skin and mucosal surfaces. To date, based on genome sequence, over 100 HPV types have been described, and these virus types vary in both their anatomical tropism and their potential to induce malignant transformation (10). The HPV replication cycle is intimately linked with the differentiation of the host epithelium (21). To initiate an infection, the virus requires access, typically via a cut or abrasion, to basal cells where the virus maintains itself at a low copy number. Subsequently, infected cells undergo differentiation and migrate to the epithelial surface, which initiates the productive phase of virus infection, signified by genome amplification, the synthesis of structural proteins, and assembly of infectious progeny. These events are orchestrated by both transcriptional and posttranscriptional mechanisms and consequently ensure that structural proteins are only synthesized in terminally differentiated cells (61).High-level expression of the E4 protein accompanies the productive phase of the HPV life cycle (40). E4 is translated from spliced E1 E4 transcripts and encodes the first 5 amino acids from the N terminus of the E1 protein fused to the E4 coding sequence. The E4 open reading frame (ORF) is the most divergent ORF within the HPV family. While there is sequence homology between E1 E4 proteins, this is generally restricted to virus types ...

Section: Discussionmentioning

confidence: 99%

The E1^E4 Protein of Human Papillomavirus Interacts with the Serine-Arginine-Specific Protein Kinase SRPK1

Bell

Martin

Roberts

2007

“…The ONM is a continuation of the endoplasmic reticulum, whereas the INM has a unique composition and contains specific resident proteins, including the lamin B receptor, the LEM-domain proteins emerin, MAN1, and lamin-associated polypeptides (LAPs), and nurim (6,10,11). Many more INM proteins have recently been identified (45,46).…”

mentioning

confidence: 99%

Herpes Simplex Virus Infection Induces Phosphorylation and Delocalization of Emerin, a Key Inner Nuclear Membrane Protein

Morris

Hofemeister

O’Hare

2007

The inner nuclear membrane (INM) contains specialized membrane proteins that selectively interact with nuclear components including the lamina, chromatin, and DNA. Alterations in the organization of and interactions with INM and lamina components are likely to play important roles in herpesvirus replication and, in particular, exit from the nucleus. Emerin, a member of the LEM domain class of INM proteins, binds a number of nuclear components including lamins, the DNA-bridging protein BAF, and F-actin and is thought to be involved in maintaining nuclear integrity. Here we report that emerin is quantitatively modified during herpes simplex virus (HSV) infection. Modification begins early in infection, involves multiple steps, and is reversed by phosphatase treatment. Emerin phosphorylation during infection involves one or more cellular kinases but can also be influenced by the US3 viral kinase, a protein whose function is known to be involved in HSV nuclear egress. The results from biochemical extraction analyses and from immunofluorescence of the detergent-resistant population demonstrate that emerin association with the INM significantly reduced during infection. We propose that the induction of emerin phosphorylation in infected cells may be involved in nuclear egress and uncoupling interactions with targets such as the lamina, chromatin, or cytoskeletal components.The nuclear envelope is composed of a double lipid bilayer, the inner and outer nuclear membranes (INM and ONM), and is underpinned on the nucleoplasmic side by the nuclear lamina, a dense meshwork of intermediate filaments formed from interlaced dimers of the lamins A/C and B. Herpes simplex virus (HSV), like all herpesviruses, replicates and packages its genome into newly formed capsids inside the nucleus of infected cells. Progeny nucleocapsids, with a size of 100 nm, are too large to pass through nuclear pores, which have a gating mechanism for soluble proteins and assemblies through an aqueous channel with a diameter of about 9 nm (reviewed in references 1, 42, 55, and 56). The mechanism by which HSV exits the nucleus remains a matter of controversy, but it has been generally accepted that a primary pathway of exit is via nucleocapsid attachment to the INM and subsequent budding into the lumenal space, thereby acquiring a primary lipid envelope (reviewed in references 13, 32, and 51). However, we currently have limited understanding of the modifications to the INM and lamina, the interactions between these components, and the mechanism involved in different stages during exit from the nucleus.The ONM is a continuation of the endoplasmic reticulum, whereas the INM has a unique composition and contains specific resident proteins, including the lamin B receptor, the LEM-domain proteins emerin, MAN1, and lamin-associated polypeptides (LAPs), and nurim (6, 10, 11). Many more INM proteins have recently been identified (45,46). Although the precise routes of localization are not known in all cases, one mechanism by which these proteins are thought to ...

“…The outer nuclear membrane is a continuation of the endoplasmic reticulum (ER), while the INM has a unique composition and contains specific resident proteins, including lamin B receptor (LBR), lamin-associated polypeptides (LAPs), emerin, and nurim (reviewed by Chu et al. [10] and Dechat et al [13]). While there are exceptions, many of these proteins are thought to localize to the INM by a diffusion-and-retention mechanism.…”

mentioning

confidence: 99%

Fate of the Inner Nuclear Membrane Protein Lamin B Receptor and Nuclear Lamins in Herpes Simplex Virus Type 1 Infection

Scott

O’Hare

2001

112

During herpesvirus egress, capsids bud through the inner nuclear membrane. Underlying this membrane is the nuclear lamina, a meshwork of intermediate filaments with which it is tightly associated. Details of alterations to the lamina and the inner nuclear membrane during infection and the mechanisms involved in capsid transport across these structures remain unclear. Here we describe the fate of key protein components of the nuclear envelope and lamina during herpes simplex virus type 1 (HSV-1) infection. We followed the distribution of the inner nuclear membrane protein lamin B receptor (LBR) and lamins A and B(2) tagged with green fluorescent protein (GFP) in live infected cells. Together with additional results from indirect immunofluorescence, our studies reveal major morphologic distortion of nuclear-rim LBR and lamins A/C, B(1), and B(2). By 8 h p.i., we also observed a significant redistribution of LBR-GFP to the endoplasmic reticulum, where it colocalized with a subpopulation of cytoplasmic glycoprotein B by immunofluorescence. In addition, analysis by fluorescence recovery after photobleaching reveals that LBR-GFP exhibited increased diffusional mobility within the nuclear membrane of infected cells. This is consistent with the disruption of interactions between LBR and the underlying lamina. In addition to studying stably expressed GFP-lamins by fluorescence microscopy, we studied endogenous A- and B-type lamins in infected cells by Western blotting. Both approaches reveal a loss of lamins associated with virus infection. These data indicate major disruption of the nuclear envelope and lamina of HSV-1-infected cells and are consistent with a virus-induced dismantling of the nuclear lamina, possibly in order to gain access to the inner nuclear membrane.