Cell surface expression of 5-hydroxytryptamine type 3 receptors is controlled by an endoplasmic reticulum retention signal.
Two subunits of the 5-hydroxytryptamine type 3 (5-HT3) have been identified (5-HT3A and 5-HT3B) that assemble into homomeric (5-HT3A) and heteromeric (5-HT3A+5-HT3B) complexes. Unassembled 5-HT3B subunits are efficiently retained within the cell. In this study, we address the mechanism controlling the release of 5-HT3B from the endoplasmic reticulum (ER). An analysis of chimeric 5-HT3A receptor(R).5-HT3BR constructs suggests the presence of elements downstream of the first transmembrane domain of 5-HT3B subunits that inhibit cell surface expression. To investigate this possibility, truncated 5-HT3B subunits were constructed and assessed for their ability to access the cell surface in COS-7 and ts201 cells. Using this approach, we have identified the presence of an ER retention signal located within the first cytoplasmic loop between transmembrane domains I and II of 5-HT3B. Transplantation of this signal (CRAR) into the homologous region of 5-HT3A results in the ER retention of this subunit until rescued by co-assembly with wild-type 5-HT3A. The mutation of this ER retention signal in 5-HT3B (5-HT3BSGER) does not lead to cell surface expression, suggesting the presence of other signals or mechanisms to control the surface expression of 5-HT3BRs. The generation of truncated 5-HT3BSGER constructs confirmed that the CRAR signal does play an important role in the ER retention of 5-HT3B.
Department of Pharmacology and Neuroscience, Ninewells Medical School, University of Dundee, Dundee DD1 9SY, Scotland.
Gary W
GW
Boyd
Anne I
AI
Doward
Ewen F
EF
Kirkness
Neil S
NS
Millar
Christopher N
CN
Connolly
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006801:Humans; D008856:Microscopy, Fluorescence; D017434:Protein Structure, Tertiary; D011985:Receptors, Serotonin / Q000096:biosynthesis* / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction
D018076:DNA, Complementary; D011985:Receptors, Serotonin; D011993:Recombinant Fusion Proteins; D004247:DNA