Role of N-Linked Glycosylation of the 5-HT
            <sub>2A</sub>
            Receptor in JC Virus Infection

Maginnis, Melissa S.; Haley, Sheila A.; Gee, Gretchen V.; Atwood, Walter J.

doi:10.1128/jvi.00978-10

Cited by 49 publications

(46 citation statements)

References 44 publications

(50 reference statements)

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“…1C). As previously shown, the fusion of the YFP tag to the C terminus did not affect viral infection (35).…”

Section: Methodssupporting

confidence: 51%

“…Given that all members of the 5-HT 2 receptor family can support JCPyV infection, it is possible that the 5-HT receptors serve a redundant role in JCPyV infection, or they may functionally cooperate to facilitate JCPyV entry. In fact, HEK293A cells express a very low level of these receptors (data not shown) and have been previously used as model to study 5-HT 2A R by exogenous overexpression (35,64). PML is a severely debilitating disease, and if left untreated, it is rapidly fatal.…”

Section: Discussionmentioning

confidence: 99%

“…The cDNAs for human 5-HT 1A R, 5-HT 1B R, 5-HT 1D R, 5-HT 1E R, 5-HT 1F R, 5-HT 2A R, 5-HT 2B R, 5-HT 2C R, 5-HT 3 R, 5-HT 4 R, 5-HT 5 R, 5-HT 6 R, 5-HT 7A R, and 5-HT 7B R in pcDNA3.1 were purchased from the Missouri S&T cDNA Resource Center (www.cdna.org). -The generation of the 5-HT 2A R-YFP (YFP stands for yellow fusion protein) fusion construct was previously described (35). To generate the 5-HT 2B R-YFP and 5-HT 2C R-YFP fusion constructs, the cDNA for both human 5-HT 2B R and 5-HT 2C R in pcDNA3.1 was PCR amplified with a 5= primer containing an XhoI site followed by the Kozak sequence and the first 16 Stable transfection of HEK293A cells.…”

Section: Methodsmentioning

confidence: 99%

“…However, the mechanism by which 5-HT 2A R contributes to JCPyV infection is not fully understood. Mutational analysis of the glycosylation sites on the N terminus of 5-HT 2A R revealed that LSTc is not linked to the receptor (35). Conversely, it has been reported that JCPyV infection can occur in the absence of 5-HT 2A R in human brain microvascular endothelial cells (36).…”

mentioning

confidence: 99%

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5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

Assetta

Maginnis

Ahufinger

et al. 2013

J Virol

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“…1C). As previously shown, the fusion of the YFP tag to the C terminus did not affect viral infection (35).…”

Section: Methodssupporting

confidence: 51%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

Assetta

Maginnis

Ahufinger

et al. 2013

J Virol

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“…It is known that the 5-HT 2 family of receptors is the functional receptor for JCPyV entry into cells. 38,39,50 All three 5-HT 2 family members colocalized with MUC1 on the collecting ducts and distal tubules of the kidney (Figure 2, A, C, and E). 5-HT 2A appears specifically on the apical side of the tubules, whereas 5-HT 2B and 5-HT 2C also label the basolateral sides.…”

Section: Jcpyv Specifically Binds To the Distal Tubules And Collectinmentioning

confidence: 95%

Human Polyomavirus Receptor Distribution in Brain Parenchyma Contrasts with Receptor Distribution in Kidney and Choroid Plexus

Haley

O’Hara

Nelson

et al. 2015

The American Journal of Pathology

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The human polyomavirus, JCPyV, is the causative agent of progressive multifocal leukoencephalopathy, a rare demyelinating disease that occurs in the setting of prolonged immunosuppression. After initial asymptomatic infection, the virus establishes lifelong persistence in the kidney and possibly other extraneural sites. In rare instances, the virus traffics to the central nervous system, where oligodendrocytes, astrocytes, and glial precursors are susceptible to lytic infection, resulting in progressive multifocal leukoencephalopathy. The mechanisms by which the virus traffics to the central nervous system from peripheral sites remain unknown. Lactoseries tetrasaccharide c (LSTc), a pentasaccharide containing a terminal a2,6elinked sialic acid, is the major attachment receptor for polyomavirus. In addition to LSTc, type 2 serotonin receptors are required for facilitating virus entry into susceptible cells. We studied the distribution of virus receptors in kidney and brain using lectins, antibodies, and labeled virus. The distribution of LSTc, serotonin receptors, and virus binding sites overlapped in kidney and in the choroid plexus. In brain parenchyma, serotonin receptors were expressed on oligodendrocytes and astrocytes, but these cells were negative for LSTc and did not bind virus. LSTc was instead found on microglia and vascular endothelium, to which virus bound abundantly. Receptor distribution was not changed in the brains of patients with progressive multifocal leukoencephalopathy. Virus infection of oligodendrocytes and astrocytes during disease progression is LSTc independent. (Am J Pathol 2015, 185: 2246e2258; http://dx.doi.org/10.1016/j.ajpath.2015.04.003)The human polyomavirus (JCPyV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a rapidly progressing, often fatal neurodegenerative disease. Although PML is rare, JCPyV infection is widespread, infecting approximately 50% to 80% of the healthy adult population. 1,2 As the initial infection is asymptomatic, the mode of JCPyV transmission is unknown. The virus establishes a persistent infection in the kidney and urinary tract of immunocompetent hosts, 3 and about 20% of these infected individuals shed virus in their urine. 4 JCPyV DNA has also been detected in other tissues, including B lymphocytes in the bone marrow, tonsillar stromal cells, lungs, spleen, and brain, 5e13 suggesting additional sites of viral persistence. The route of viral transmission from the initial site(s) of infection and latency to the central nervous system (CNS), the main site of pathogenesis, is not clearly understood.Under conditions of immunosuppression, JCPyV infects and destroys the myelin-producing oligodendrocytes, resulting in demyelination, which is the hallmark of this fatal disease; to a Supported by NIH grants R01NS043097 (W.J.A.) and P01NS065719 (W.J.A.).

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Comprehensive structural modeling and preparation of human 5‐HT_2A G‐protein coupled receptor in functionally active form

et al. 2019

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The serotonin 2A receptor (5-HT 2A R) is an important member of the G-protein coupled receptor (GPCR) family involved in an array of neuromodulatory functions. Although the high-resolution structures of truncated versions of GPCRs, captured in ligand-bound conformational states, are available, the structures lack several functional regions, which have crucial roles in receptor response. Here, in order to understand the structure and dynamics of the ligand-free form of the receptor, we have performed meticulous modeling of the 5-HT 2A R with the third intracellular loop (ICL3). Our analyses revealed that the ligand-free ground state structure of 5-HT 2A R has marked distinction with ligand-bound conformations of 5-HT 2 subfamily proteins and exhibits extensive backbone flexibility across the loop regions, suggesting the importance of purifying the receptor in its native form for further studies. Hence, we have standardized a strategy that efficiently increases the expression of 5-HT 2A R by infecting Sf9 cells with a very low multiplicity of infection of baculovirus in conjunction with production boost additive and subsequently, purify the full-length receptor. Furthermore, we have optimized the selective over-expression of glycosylated and nonglycosylated forms of the receptor merely by switching the postinfection growth time, a method that has not been reported earlier. K E Y W O R D S 5-HT 2A GPCR, baculoviral expression system, homology modeling, molecular dynamics simulation, protein over-expression and purification

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Role of N-Linked Glycosylation of the 5-HT _2A Receptor in JC Virus Infection

Cited by 49 publications

References 44 publications

5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

Human Polyomavirus Receptor Distribution in Brain Parenchyma Contrasts with Receptor Distribution in Kidney and Choroid Plexus

Comprehensive structural modeling and preparation of human 5‐HT_2A G‐protein coupled receptor in functionally active form

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Role of N-Linked Glycosylation of the 5-HT 2A Receptor in JC Virus Infection

Cited by 49 publications

References 44 publications

5-HT 2 Receptors Facilitate JC Polyomavirus Entry

5-HT 2 Receptors Facilitate JC Polyomavirus Entry

Human Polyomavirus Receptor Distribution in Brain Parenchyma Contrasts with Receptor Distribution in Kidney and Choroid Plexus

Comprehensive structural modeling and preparation of human 5‐HT2A G‐protein coupled receptor in functionally active form

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Product

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Role of N-Linked Glycosylation of the 5-HT _2A Receptor in JC Virus Infection

5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

5-HT ₂ Receptors Facilitate JC Polyomavirus Entry

Comprehensive structural modeling and preparation of human 5‐HT_2A G‐protein coupled receptor in functionally active form