Structural analysis of a MIP family protein from the digestive tract of Cicadella viridis.
Homopteran insects, and especially Cicadella viridis, display in their digestive tract a specialized epithelial differentiation, the filter chamber (FC) acting as a water-shunting complex. The main intrinsic membrane protein of the FC is a 25,000-Da polypeptide (P25). In this paper we demonstrate that this P25 polypeptide is a member of the MIP family of membrane channel proteins, and that P25 forms homotetramers in the native membranes. Using polymerase chain reaction, a 360-base pair cDNA, named cic, was isolated from RNA of the FC. cic encodes a 119-amino acid polypeptide (CIC) whose homologies with MIP26, AQP1 (CHIP), AQP2, and gamma-TIP are 38, 38, 34, and 20%, respectively. Using a specific antibody raised against a 15-amino acid peptide from the CIC sequence, we concluded that CIC and P25 are identical entities, and hence that P25 belongs to the MIP family. We investigated the quaternary structure of P25 in the membranes of the FC using biophysical analysis of P25 nondenaturing detergent micelles, scanning transmission electron microscopy, and image processing of conventional transmission electron microscopic images. All those different approaches converged to the conclusion that P25 exists as an homotetramer forming a regular two-dimensional array in the membranes.
Beuron
F
F
Laboratoire de Biologie Cellulaire, URA CNRS No 256, Université de Rennes 1, France.
Le Cahérec
F
F
Guillam
M T
MT
Cavalier
A
A
Garret
A
A
Tassan
J P
JP
Delamarche
C
C
Schultz
P
P
Mallouh
V
V
Rolland
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins; D001483:Base Sequence; D005136:Eye Proteins / Q000737:chemistry* / Q000235:genetics; D007313:Insecta / Q000737:chemistry*; D007473:Ion Channels / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis
D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins; D005136:Eye Proteins; D007473:Ion Channels; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; C030328:aquaporin 0