Novel shuttling domain in a regulator (RSC1A1) of transporter SGLT1 steers cell cycle-dependent nuclear location.
The gene product of RSC1A1, RS1, participates in the regulation of the Na(+)-D-glucose cotransporter SGLT1. RS1 inhibits release of SGLT1 from the trans Golgi network. In subconfluent LLC-PK(1) cells, RS1 migrates into the nucleus and modulates transcription of SGLT1, whereas most confluent cells do not contain RS1 in the nuclei. We showed that confluence-dependent nuclear location of RS1 is because of different phases of the cell cycle and identified a RS1 nuclear shuttling domain (RNS) with an associated protein kinase C (PKC) phosphorylation site (RNS-PKC) that mediates cell cycle-dependent nuclear location. RNS-PKC contains a novel non-conventional nuclear localization signal interacting with importin beta1, a nuclear export signal mediating export via protein CRM1 and a Ca(2+)-dependent calmodulin binding site. PKC and calmodulin compete for binding to RNS-PKC. Mutagenesis experiments and analyses of the phosphorylation status suggest the following sequences of events. Subconfluent cells without and with synchronization to the G2/M phase contain non-phosphorylated RNS-PKC that mediates nuclear import of RS1 but not its export. During confluence or synchronization of subconfluent cells to the G2/M phase, phosphorylation of RNS-PKC mediates rapid nuclear export of RS1.
Filatova
Alina
A
Institute of Anatomy and Cell Biology, University of Würzburg, 97070 Würzburg, Germany.
Leyerer
Marina
M
Gorboulev
Valentin
V
Chintalapati
Chakravarthi
C
Reinders
Yvonne
Y
Müller
Thomas D
TD
Srinivasan
Aruna
A
Hübner
Stefan
S
Koepsell
Hermann
H
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002453:Cell Cycle; D002467:Cell Nucleus / Q000378:metabolism*; D016895:Culture Media, Serum-Free; D005822:Genetic Vectors; D005947:Glucose / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D028884:Karyopherins / Q000378:metabolism; D018374:LLC-PK1 Cells; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019913:Nuclear Localization Signals / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D051269:Sodium-Glucose Transporter 1 / Q000378:metabolism*; D013552:Swine; D013997:Time Factors; D014162:Transfection; D021601:trans-Golgi Network / Q000378:metabolism
D016895:Culture Media, Serum-Free; D028884:Karyopherins; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D017411:Ribonucleoproteins, Small Nuclear; D051269:Sodium-Glucose Transporter 1; D049452:Green Fluorescent Proteins; D012964:Sodium; D011493:Protein Kinase C; D005947:Glucose