Retinitis pigmentosa (RP)1
is an inherited retinal disease affecting approximately one in 3,500 individuals [1
]. The progressive vision loss results from apoptosis of rod and cone photoreceptor cells [2
]. RP is often caused by mutations in photoreceptor-specific proteins, but can also result from mutations in widely expressed proteins such as IMPDH1 [4
]. The most commonly occurring IMPDH1 mutation, D226N, accounts for 1–2.5% of all autosomal dominant RP cases [6
]. Mutations in IMPDH1 also cause Leber congenital amaurosis (LCA), a more severe retinal degeneration [7
]. The pathophysiological mechanism of IMPDH1-linked adRP and the tissue specificity of disease are not understood.
IMPDH catalyzes the controlling step in de novo
synthesis of guanine nucleotide. Mammals express two isozymes, IMPDH1 and IMPDH2, which are 84% identical. To the first approximation, IMPDH1 is constitutively expressed and IMPDH2 is amplified in proliferating tissues [9
]. IMPDH1 predominates in the retina [11
It is unlikely that adRP results from a loss of IMPDH1 function in the retina. First, the adRP-linked mutations of IMPDH1 have no effect on enzymatic activity [7
]. Moreover, most of the mutations are located in a subdomain that is not required for enzymatic function [13
]. Second, drugs targeting IMPDH are widely used as immunosuppressive, anticancer and antiviral therapy, yet no visual side effects have been reported. Lastly, IMPDH1 knocked-out mice display only mild retinopathy [15
]. We have discovered that IMPDH binds nucleic acids, and that this interaction is mediated by the subdomain [16
]. The adRP-linked mutations perturb this nucleic binding function, decreasing both affinity and specificity of this interaction [7
We have recently discovered that human retina contains two novel isoforms of IMPDH1, IMPDH1(546) (major) and IMPDH1(595) (minor) derived from alternative mRNA splicing [these proteins are also known as IMPDH1α/IMPDH1(13b) and IMPDHγ/IMPDH1(A+13b), respectively] [11
]. Both IMPDH1(546) and IMPDH1(595) contain a 32 residue C-terminal extension while IMPDH1(595) has an additional 49 residues on the N-terminus. Since poly-His and GFP tags do not perturb either the enzymatic or nucleic acid binding actitvities of IMPDH [12
], these N- and C-terminal extensions need not perturb function. The retinal-specific expression of IMPDH1 isoforms and the compensating presence of IMPDH2 in other tissues may account for the photoreceptor-specific effects of the adRP/LCA-causing mutations, but do not explain the pathological mechanism.
Here we characterize the two retinal isoforms of IMPDH1 as well as their D226N variants. The enzymatic activity of these isoforms is indistinguishable from the canonical IMPDH1. In contrast, neither retinal isoform binds nucleic acids. This observation suggests that C-terminal extension blocks the nucleic acid binding site, perhaps by interacting with the subdomain. We propose that photoreceptors contain additional proteins that interact with the C-terminal extensions to regulate the nucleic acid binding properties of the retinal isoforms.