Serine 13 of the human cytomegalovirus viral cyclin-dependent kinase UL97 is required for regulatory protein 14-3-3 binding and UL97 stability

Iwahori, Satoko; Umaña, Angie C.; Kalejta, Robert F.; Murata, Takayuki

doi:10.1016/j.jbc.2022.102513

“…An important report was recently published addressing the role of phosphorylation on CHPK and stability. Iwahori et al ., [78] showed that binding of the regulatory proteins in the 14-3-3 family were important for stability of the human cytomegalovirus CHPK (cyclin-dependent kinase, vCDK UL97). We have consistently seen stability issues with MDV pCHPK in both cell culture and in chickens [23] when its kinase activity is abrogated (vCHPKmut).…”

Section: Discussionmentioning

confidence: 99%

The alphaherpesvirus conserved pUS10 is important for natural infection and its expression is regulated by the conservedHerpesviridaeprotein kinase (CHPK)

Ponnuraj

¹

,

Akbar

²

,

Arrington

³

et al. 2022

Preprint

0

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Conserved Herpesviridae protein kinases (CHPK) are conserved among all members of the Herpesviridae. Herpesviruses lacking CHPK propagate in cell culture at varying degrees, depending on the virus and cell culture system. CHPK is dispensable for Marek’s disease herpesvirus (MDV) replication in cell culture and experimental infection in chickens; however, CHPK—particularly its kinase activity—is essential for horizontal transmission in chickens, also known as natural infection. To address the importance of CHPK during natural infection in chickens, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) based proteomics of samples collected from live chickens. Comparing modification of viral proteins in feather follicle epithelial (FFE) cells infected with wildtype or a CHPK-null virus, we identified the US10 protein (pUS10) as a potential target for CHPK in vivo. When expression of pUS10 was evaluated in cell culture and in FFE skin cells during in vivo infection, pUS10 was severely reduced or abrogated in cells infected with CHPK mutant or CHPK-null viruses, respectively, indicating a potential role for pUS10 in transmission. To test this hypothesis, US10 was deleted from the MDV genome, and the reconstituted virus was tested for replication, horizontal transmission, and disease induction. Our results showed that removal of US10 had no effect on the ability of MDV to transmit in experimentally infected chickens, but disease induction in naturally infected chickens was significantly reduced. These results show CHPK is necessary for pUS10 expression both in cell culture and in the host, and pUS10 is important for disease induction during natural infection.

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“…An important report was recently published addressing the role of phosphorylation on CHPK and stability. Iwahori et al, [78] showed that binding of the regulatory proteins in the 14-3-3 family were important for stability of the human cytomegalovirus CHPK (cyclindependent kinase, vCDK UL97). We have consistently seen stability issues with MDV pCHPK in both cell culture and in chickens [23] when its kinase activity is abrogated (vCHPKmut).…”

Section: Plos Pathogensmentioning

confidence: 99%

The alphaherpesvirus conserved pUS10 is important for natural infection and its expression is regulated by the conserved Herpesviridae protein kinase (CHPK)

Ponnuraj

¹

,

Akbar

²

,

Arrington

³

et al. 2023

View full text Add to dashboard Cite

Conserved Herpesviridae protein kinases (CHPK) are conserved among all members of the Herpesviridae. Herpesviruses lacking CHPK propagate in cell culture at varying degrees, depending on the virus and cell culture system. CHPK is dispensable for Marek’s disease herpesvirus (MDV) replication in cell culture and experimental infection in chickens; however, CHPK—particularly its kinase activity—is essential for horizontal transmission in chickens, also known as natural infection. To address the importance of CHPK during natural infection in chickens, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) based proteomics of samples collected from live chickens. Comparing modification of viral proteins in feather follicle epithelial (FFE) cells infected with wildtype or a CHPK-null virus, we identified the US10 protein (pUS10) as a potential target for CHPK in vivo. When expression of pUS10 was evaluated in cell culture and in FFE skin cells during in vivo infection, pUS10 was severely reduced or abrogated in cells infected with CHPK mutant or CHPK-null viruses, respectively, indicating a potential role for pUS10 in transmission. To test this hypothesis, US10 was deleted from the MDV genome, and the reconstituted virus was tested for replication, horizontal transmission, and disease induction. Our results showed that removal of US10 had no effect on the ability of MDV to transmit in experimentally infected chickens, but disease induction in naturally infected chickens was significantly reduced. These results show CHPK is necessary for pUS10 expression both in cell culture and in the host, and pUS10 is important for disease induction during natural infection.

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