Single mutations in the S4 domain of cone photoreceptor cGMP-gated channels, characteristic of human achromatopsia, cause functional channel failure

FAILLACE MP, BERNABEU RO, KORENBROT JI

Fort Lauderdale, Florida, USA

Congreso; Association for Research in Vision and Ophtalmology Annual Meeting 2004; 2004

ARVO - Association for Research in Vision and Ophtalmology

Purpose: cGMP-gated ion channels in retinal photoreceptors (CNG channels) are activated by cGMP, but are not voltage-dependent. Yet, their structure includes an S4 motif consisting of tandem repeats of the R(or K)XX sequence within the 4th transmembrane helix. S4 is characteristic of every known voltage-gated ion channel and mutations in S4 in these channels alter their voltage-dependent function. Human achromatopsia is a congenital loss of cone function and a frequent mutation in this patient population is a single amino acid substitution in S4 of cone CNGA3 channels. We investigated the functional consequence of point mutations in S4 of bovine CNGA3 cone channels by assessing the electrical properties and cell protein processing pattern of transmembrane helix. Results: In a mammalian cell line, wild type bovine cone photoreceptor channel alpha subunits (bCNGA3) convert from an unglycosylated state, at 90kDa, to two glycosylated states at 93 and 102 kDa as they transit within the cell to their final location at the plasma membrane. Glycosylation per-se is not required to yield functional channels, yet it is a milestone that distinguishes sequential steps in channel protein maturation. Point mutation of specific, not all, charged and neutral residues within S4 cause failure of functional channel expression.  Cellular channel protein processing fails in every one of the non-functional S4 mutations we studied.  Mutant proteins do not reach the 102 kDa glycosylated state and do not arrive at the plasma membrane.  They remain trapped within the endoplasmic reticulum and fail to transit out to the Golgi apparatus. Coexpression of cone CNG beta subunit (CNGB3) does not rescue the consequence of S4 mutations in CNGA3. Conclusions:   Point mutations in the S4 cone CNGA3 channel prevent proper protein trafficking within cells. This class of mutations occurs in achromatopsia and probably causes the loss of cone function characteristic of this congenital disease. Single mutations in the S4 domain of cone photoreceptor cGMP-gated channels, characteristic of human achromatopsia, cause functional channel failure. It is likely that an intact S4 is required for proper protein folding and/or assembly in the endoplasmic reticulum membrane.