The nop-1 gene of Neurospora crassa encodes a seven transmembrane helix retinal-binding protein homologous to archaeal rhodopsins

Abstract

Opsins are a class of retinal-binding, seven transmembrane helix proteins that function as lightresponsive ion pumps or sensory receptors. Previously, genes encoding opsins had been identified in animals and the Archaea but not in fungi or other eukaryotic microorganisms. Here, we report the identification and mutational analysis of an opsin gene, nop-1, from the eukaryotic filamentous fungus Neurospora crassa. The nop-1 amino acid sequence predicts a protein that shares up to 81.8% amino acid identity with archaeal opsins in the 22 retinal binding pocket residues, including the conserved lysine residue that forms a Schiff base linkage with retinal. Evolutionary analysis revealed relatedness not only between NOP-1 and archaeal opsins but also between NOP-1 and several fungal opsin-related proteins that lack the Schiff base lysine residue. The results provide evidence for a eukaryotic opsin family homologous to the archaeal opsins, providing a plausible link between archaeal and visual opsins. Extensive analysis of ⌬nop-1 strains did not reveal obvious defects in light-regulated processes under normal laboratory conditions. However, results from Northern analysis support light and conidiation-based regulation of nop-1 gene expression, and NOP-1 protein heterologously expressed in Pichia pastoris is labeled by using all-trans [ 3 H]retinal, suggesting that NOP-1 functions as a rhodopsin in N. crassa photobiology.Retinal is a chromophore that binds to integral membrane proteins (opsins) to form light-absorbing pigments known as rhodopsins. Visual rhodopsins contain the chromophore 11-cis retinal, which has intrinsic qualities optimal for vision (1). In contrast, archaeal rhodopsins contain all-trans retinal and function in light-activated ion pumping and phototaxis (reviewed in ref. 2). Archaeal and visual rhodopsins show little sequence identity but possess a similar secondary structure (3). This structure includes seven transmembrane ␣-helical domains and a conserved lysine residue in the seventh helix (helix G) that forms a Schiff base linkage with retinal (4). On light absorption, the retinal isomerizes and the Schiff base is deprotonated. These events are followed by conformational changes in the opsin protein and subsequent transduction of the light signal.Four different archaeal rhodopsins have been identified in Halobacterium salinarum (reviewed in refs. 2 and 5). These are bacteriorhodopsin, (BR), halorhodopsin (HR), Sensory rhodopsin I (SRI), and sensory rhodopsin II (SRII). BR and HR are ion pumps that are activated by orange light under semianaerobic conditions; they function by expelling hydrogen ions from the cell or taking in extracellular chloride ions, respectively. The pumping action of BR and HR hyperpolarizes the membrane, driving ATP synthesis during anaerobic growth. SRI controls phototaxis of the cell to orange light during anaerobic conditions to energize pumping of BR and HR. SRII also controls cell motility but is produced only in oxygen, where it induces an avoidance response to b...

Lessons from the Genome Sequence ofNeurospora crassa: Tracing the Path from Genomic Blueprint to Multicellular Organism

“…These proteins fall into five families: microbial opsins, pheromone receptors, glucose sensors, nitrogen sensors, and a novel class, not previously identified in fungi. The first class consists of an opsin (nop-1) and an opsin-related protein (orp-1) that were identified during prior EST projects (78); this family has three members in S. cerevisiae (Hsp30p, Yro2p and Mrh1p) (78,297,592,861). In the second family are two predicted pheromone receptors, similar to S. cerevisiae ␣-factor and a-factor pheromone receptors (reviewed in reference 199).…”

Leptosphaeria rhodopsin: Bacteriorhodopsin-like proton pump from a eukaryote

“…Microbial rhodopsins bind all-trans-retinal (as opposed to 11-cis-retinal-based visual pigments) and have two primary functions, photosensory transduction and light-driven ion transport (3). To date, the ion transport function has only been observed in the archaea and eubacteria, with the function of eukaryotic rhodopsins of haloarchaeal type suggested to be photosensory, as in the fungi Allomyces reticulatus and Neurospora crassa (4)(5)(6). Recently, the photosensory function was demonstrated for the rhodopsins of the alga Chlamydomonas reinhardtii (7), where rhodopsin constitutes a light-sensitive domain in a larger membrane protein (7,8).…”

brp and blh Are Required for Synthesis of the Retinal Cofactor of Bacteriorhodopsin in Halobacterium salinarum

“…Two distinct rhodopsin families are known: the visual rhodopsins, which bind 11-cis retinal or related compounds and function as photoreceptors in vertebrates (1) and invertebrates (2), and the archaeal rhodopsins, which bind all-trans-retinal and function as light-driven ion pumps and phototaxis receptors in archaea (3). Archaeal rhodopsin orthologs have been found recently in bacteria (4) and fungi (5), suggesting that retinal-based pigments are of widespread significance. Despite their importance, the biogenesis of these molecules is not fully understood.…”