title: Knockout of GARPs and the β-subunit of the rod cGMP-gated channel disrupts disk morphogenesis and rod outer segment structural integrity.
abstract: Ion flow into the rod photoreceptor outer segment (ROS) is regulated by a member of the cyclic-nucleotide-gated cation-channel family; this channel consists of two subunit types, alpha and beta. In the rod cells, the Cngb1 locus encodes the channel beta-subunit and two related glutamic-acid-rich proteins (GARPs). Despite intensive research, it is still unclear why the beta-subunit and GARPs are coexpressed and what function these proteins serve. We hypothesized a role for the proteins in the maintenance of ROS structural integrity. To test this hypothesis, we created a Cngb1 5'-knockout photoreceptor null (Cngb1-X1). Morphologically, ROSs were shorter and, in most rods that were examined, some disks were misaligned, misshapen and abnormally elongated at periods when stratification was still apparent and degeneration was limited. Additionally, a marked reduction in the level of channel alpha-subunit, guanylate cyclase I (GC1) and ATP-binding cassette transporter (ABCA4) was observed without affecting levels of other ROS proteins, consistent with a requirement for the beta-subunit in channel assembly or targeting of select proteins to ROS. Remarkably, phototransduction still occurred when only trace levels of homomeric alpha-subunit channels were present, although rod sensitivity and response amplitude were both substantially reduced. Our results demonstrate that the beta-subunit and GARPs are necessary not only to maintain ROS structural integrity but also for normal disk morphogenesis, and that the beta-subunit is required for normal light sensitivity of the rods.
authors.affiliation: Department of Vision Sciences, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
authors.forename: Youwen
authors.initials: Y
authors.lastname: Zhang
authors.forename: Laurie L
authors.initials: LL
authors.lastname: Molday
authors.forename: Robert S
authors.initials: RS
authors.lastname: Molday
authors.forename: Shanta S
authors.initials: SS
authors.lastname: Sarfare
authors.forename: Michael L
authors.initials: ML
authors.lastname: Woodruff
authors.forename: Gordon L
authors.initials: GL
authors.lastname: Fain
authors.forename: Timothy W
authors.initials: TW
authors.lastname: Kraft
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Pittler