The product of C9orf72, a gene strongly implicated in neurodegeneration, is structurally related to DENN Rab-GEFs

Abstract

Motivation: Fronto-temporal dementia (FTD) and amyotrophic lateral sclerosis (ALS, also called motor neuron disease, MND) are severe neurodegenerative diseases that show considerable overlap at the clinical and cellular level. The most common single mutation in families with FTD or ALS has recently been mapped to a non-coding repeat expansion in the uncharacterized gene C9ORF72. Although a plausible mechanism for disease is that aberrant C9ORF72 mRNA poisons splicing, it is important to determine the cellular function of C9ORF72, about which nothing is known.

Results: Sensitive homology searches showed that C9ORF72 is a full-length distant homologue of proteins related to Differentially Expressed in Normal and Neoplasia (DENN), which is a GDP/GTP exchange factor (GEF) that activates Rab-GTPases. Our results suggest that C9ORF72 is likely to regulate membrane traffic in conjunction with Rab-GTPase switches, and we propose to name the gene and its product DENN-like 72 (DENNL72).

Fig. 1.

C9ORF72 is structurally homologous to DENN-like proteins. Probabilities of shared structure (p^SS) of pairwise comparisons of sequences by HHpred, comparing in (A): C9ORF72, FLCN, LCHN, two AVL9 sequences (human Hs, yeast Sc), DENND1B; and in (B): FLCN, SMcr8, NPRL2, DENND1B, C9ORF72, FNIP (worm). Values are the maximum p^SS obtained after optimizing searches. For details of query alignments and hits, see Supplementary Table S2. Variations: * match is to PDB entry for FLCN; ^• match is to worm C9ORF72. Self-searches all give p^SS = 100%

Fig. 2.

Relationships between C9ORF72, DENNs and other structural homologues. (A) Secondary structural conservation between DENN-like proteins and C9ORF72. Predicted helices (red) and sheets (blue, light blue = low confidence) are shown in C9ORF72, Avl9p, NPRL2 N-terminus and FNIP C-terminus. Structural elements are numbered in Avl9p according to homology to DENND1B (Wu et al., 2011). Solid arrows join elements aligned by HHpred, and other alignments were made by hand (dashed arrows). Variations in C9ORF72 are indicated by numbers above. Large brackets indicate the extent of match between C9ORF72 and AVL9 (i) initially and (ii) in the reverse search. Insertions are indicated by the number of residues, except for ‘+’, indicating a loop of 130 aa in human AVL9. (B) Cluster map of the DENN-like superfamily. 306 diverse members of 11 sub-families of DENN-like proteins, excluding LD only NPRL2/3s, were tested for all-versus-all similarity (10 iterations of PSI-BLAST), and clustered by CLANS (Frickey and Lupas, 2004). All pairs with similarity threshold P ≤ 10⁻²⁵ are joined by lines, which are darker and shorter with greater similarity. Sub-families are identified by colour. Outlier organisms (Trichomonas and Entamoeba) are identified by shape. Thirty-one colicins included as negative controls showed no links to the other sequences (data not shown)