Human serotonin transporter variants display altered sensitivity to protein kinase G and p38 mitogen-activated protein kinase

Harish C. Prasad, Chong Bin Zhu, Jacob L McCauley, Devadoss J. Samuvel, Sammanda Ramamoorthy, Richard C. Shelton, William A. Hewlett, James S. Sutcliffe, Randy D. Blakely

Research output: Contribution to journalArticle

132 Citations (Scopus)

Abstract

Human serotonin [5-hydroxytryptamine (5-HT)] transporters (hSERT, SHTT, and SLC6A4) inactivate S-HT after release and are prominent targets for therapeutic intervention in mood, anxiety, and obsessive-compulsive disorders. Multiple hSERT coding variants have been identified, although to date no comprehensive functional analysis of these variants has been reported. We transfected hSERT or 10 hSERT coding variants and examined total and surface protein expression, antagonist recognition, and transporter modulation by posttranslational, regulatory pathways. Two variants, Pro339Leu and Ile425Val, demonstrated significant changes in surface expression supporting alterations in 5-HT transport capacity (Vmax). Regardless of basal transport activity, all SERT variants displayed a capacity for rapid, phorbol ester-triggered down-regulation. Remarkably, five variants (Thr4Ala, Gly56Ala, Glu215Lys, Lys605Asn, and Pro612Ser) demonstrated no capacity for 5-HT uptake stimulation after acute protein kinase G (PKG)/p38 mitogen-activated protein kinase (MAPK) activation. Epstein-Barr virus (EBV)-transformed lymphocytes natively expressing the most common of these variants (Gly56Ala) exhibited a similar loss of 5-HT uptake stimulation by PKG p38 MAPK activators. HeLa cells transfected with the Gly56Ala variant demonstrated elevated basal phosphorylation and, unlike hSERT, could not be further phosphorylated after 8-bromo cGMP (8BrcGMP) treatments. These studies reveal cellular phenotypes associated with naturally occurring human SERT coding variants and suggest that altered transporter regulation by means of PKG/p38 MAPK-linked pathways may influence risk for disorders attributed to compromised 5-HT signaling.

Original languageEnglish
Pages (from-to)11545-11550
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number32
DOIs
StatePublished - Aug 9 2005
Externally publishedYes

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Cyclic GMP-Dependent Protein Kinases
Serotonin Plasma Membrane Transport Proteins
p38 Mitogen-Activated Protein Kinases
Serotonin
Obsessive-Compulsive Disorder
Phorbol Esters
Human Herpesvirus 4
HeLa Cells
Membrane Proteins
Down-Regulation
Anxiety
Phosphorylation
Lymphocytes
Phenotype

Keywords

  • Antidepressant
  • Autism
  • Polymorphism
  • Regulation
  • Transport

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Human serotonin transporter variants display altered sensitivity to protein kinase G and p38 mitogen-activated protein kinase. / Prasad, Harish C.; Zhu, Chong Bin; McCauley, Jacob L; Samuvel, Devadoss J.; Ramamoorthy, Sammanda; Shelton, Richard C.; Hewlett, William A.; Sutcliffe, James S.; Blakely, Randy D.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 102, No. 32, 09.08.2005, p. 11545-11550.

Research output: Contribution to journalArticle

Prasad, Harish C. ; Zhu, Chong Bin ; McCauley, Jacob L ; Samuvel, Devadoss J. ; Ramamoorthy, Sammanda ; Shelton, Richard C. ; Hewlett, William A. ; Sutcliffe, James S. ; Blakely, Randy D. / Human serotonin transporter variants display altered sensitivity to protein kinase G and p38 mitogen-activated protein kinase. In: Proceedings of the National Academy of Sciences of the United States of America. 2005 ; Vol. 102, No. 32. pp. 11545-11550.
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AU - Prasad, Harish C.

AU - Zhu, Chong Bin

AU - McCauley, Jacob L

AU - Samuvel, Devadoss J.

AU - Ramamoorthy, Sammanda

AU - Shelton, Richard C.

AU - Hewlett, William A.

AU - Sutcliffe, James S.

AU - Blakely, Randy D.

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AB - Human serotonin [5-hydroxytryptamine (5-HT)] transporters (hSERT, SHTT, and SLC6A4) inactivate S-HT after release and are prominent targets for therapeutic intervention in mood, anxiety, and obsessive-compulsive disorders. Multiple hSERT coding variants have been identified, although to date no comprehensive functional analysis of these variants has been reported. We transfected hSERT or 10 hSERT coding variants and examined total and surface protein expression, antagonist recognition, and transporter modulation by posttranslational, regulatory pathways. Two variants, Pro339Leu and Ile425Val, demonstrated significant changes in surface expression supporting alterations in 5-HT transport capacity (Vmax). Regardless of basal transport activity, all SERT variants displayed a capacity for rapid, phorbol ester-triggered down-regulation. Remarkably, five variants (Thr4Ala, Gly56Ala, Glu215Lys, Lys605Asn, and Pro612Ser) demonstrated no capacity for 5-HT uptake stimulation after acute protein kinase G (PKG)/p38 mitogen-activated protein kinase (MAPK) activation. Epstein-Barr virus (EBV)-transformed lymphocytes natively expressing the most common of these variants (Gly56Ala) exhibited a similar loss of 5-HT uptake stimulation by PKG p38 MAPK activators. HeLa cells transfected with the Gly56Ala variant demonstrated elevated basal phosphorylation and, unlike hSERT, could not be further phosphorylated after 8-bromo cGMP (8BrcGMP) treatments. These studies reveal cellular phenotypes associated with naturally occurring human SERT coding variants and suggest that altered transporter regulation by means of PKG/p38 MAPK-linked pathways may influence risk for disorders attributed to compromised 5-HT signaling.

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KW - Autism

KW - Polymorphism

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KW - Transport

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