Expression of Rab small GTPases in epithelial Caco-2 cells: Rab21 is an apically located GTP-binding protein in polarised intestinal epithelial cells.
Rab proteins belong to a subfamily of small GTP-binding protein genes of the Ras superfamily and play an important role in intracellular vesicular targeting. The presence of members of this protein family was examined in Caco-2 cells by a PCR-based strategy. Twenty-five different partial cDNA sequences were isolated, including 18 Rab protein family members. Seven novel human sequences, representing Rab2B, Rab6A', Rab6B, Rab10, Rab19B, Rab21 and Rab22A, were identified. For one clone, encoding Rab21, full-length cDNA was isolated from a Caco-2 cDNA library. Northern blot analysis showed a ubiquitous expression pattern of Rab21. To study Rab21 protein expression in Caco-2 cells, polyclonal antibodies were raised against GST-Rab21 fusion protein and characterised. The antibodies recognised Rab21 as a protein of approximately 25 kDa. Interestingly, the protein shows a general ER-like staining in nonpolarised Caco-2 cells in contrast to an apically located vesicle-like staining in polarised Caco-2 cells. Furthermore, immunohistochemical staining on human jejunal tissue showed a predominant expression of Rab21 in the epithelial cell layer with high expression levels in the apical region, whereas stem cells in the crypts were negative. We therefore suggest an alternative role for Rab21 in the regulation of vesicular transport in polarised intestinal epithelial cells.
Opdam
F J
FJ
Department of Cell Biology, Institute of Cellular Signalling, University of Nijmegen, The Netherlands.
Kamps
G
G
Croes
H
H
van Bokhoven
H
H
Ginsel
L A
LA
Fransen
J A
JA
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D019556:COS Cells; D018938:Caco-2 Cells; D016764:Cell Polarity; D003001:Cloning, Molecular; D004285:Dogs; D006801:Humans; D007150:Immunohistochemistry; D007413:Intestinal Mucosa / Q000166:cytology / Q000201:enzymology*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D020691:rab GTP-Binding Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; C415145:RAB21 protein, human; D020691:rab GTP-Binding Proteins