Subathra Poopalasundaram

The molecular mechanism for the spectral shifts between vertebrate ultraviolet- and violet-sensitive cone visual pigments

The short-wave-sensitive (SWS) visual pigments of vertebrate cone photoreceptors are divided into two classes on the basis of molecular identity, SWS1 and SWS2. Only the SWS1 class are present in mammals. The SWS1 pigments can be further subdivided into violet-sensitive (VS), with lambda(max) (the peak of maximal absorbance) values generally between 400 and 430 nm, and ultraviolet-sensitive (UVS), with a lambda(max)<380 nm. Phylogenetic evidence indicates that the ancestral pigment was UVS and that VS pigments have evolved separately from UVS pigments in the different vertebrate lineages. In this study, we have examined the mechanism of evolution of VS pigments in the mammalian lineage leading to present day ungulates (cow and pig). Amino acid sequence comparisons of the UVS pigments of teleost fish, amphibia, reptiles and rodents show that site 86 is invariably occupied by Phe but is replaced in bovine and porcine VS pigments by Tyr. Using site-directed mutagenesis of goldfish UVS opsin, we have shown that a Phe-86-->Tyr substitution is sufficient by itself to shift the lambda(max) of the goldfish pigment from a wild-type value of 360 nm to around 420 nm, and the reverse substitution of Tyr-86-Phe into bovine VS opsin produces a similar shift in the opposite direction. The substitution of this single amino acid is sufficient to account therefore for the evolution of bovine and porcine VS pigments. The replacement of Phe with polar Tyr at site 86 is consistent with the stabilization of Schiff-base protonation in VS pigments and the absence of protonation in UVS pigments.

Vision in the ultraviolet

Abstract. Sensitivity to ultraviolet light (UV) is achieved by photoreceptors in the eye that contain a class of visual pigments maximally sensitive to light at wavelengths <400 nm. It is widespread in the animal kingdom where it is used for mate choice, communication and foraging for food. UV sensitivity is not, however, a constant feature of the visual system, and in many vertebrate species, the UV-sensitive (UVS) pigment is replaced by a violet-sensitive (VS) pigment with maximal sensitivity between 410 and 435 nm. The role of protonation of the Schiff base-chromophore linkage and the mechanism for tuning of pigments into the UV is discussed in detail. Amino acid sequence analysis of vertebrate VS/UVS pigments indicates that the ancestral pigment was UVS, with loss of UV sensitivity occurring separately in mammals, amphibia and birds, and subsequently regained by a single amino acid substitution in certain bird species. In contrast, no loss of UV sensitivity has occurred in the UVS pigments of insects.

Divergent mechanisms for the tuning of shortwave sensitive visual pigments in vertebrates.

Of the four classes of vertebrate cone visual pigments, the shortwave-sensitive SWS1 class shows the shortest lambda(max) values with peaks in different species in either the violet (390-435 nm) or ultraviolet (around 365 nm) regions of the spectrum. Phylogenetic evidence indicates that the ancestral pigment was probably UV-sensitive (UVS) and that the shifts between violet and UV have occurred many times during evolution. This is supported by the different mechanisms for these shifts in different species. All visual pigments possess a chromophore linked via a Schiff base to a Lys residue in opsin protein. In violet-sensitive (VS) pigments, the Schiff base is protonated whereas in UVS pigments, it is almost certainly unprotonated. The generation of VS from ancestral UVS pigments most likely involved amino acid substitutions in the opsin protein that serve to stabilise protonation. The key residues in the opsin protein for this are at sites 86 and 90 that are adjacent to the Schiff base and the counterion at Glu113. In this review, the different molecular mechanisms for the UV or violet shifts are presented and discussed in the context of the structural model of bovine rhodopsin.

Neuronal DnaJ proteins HSJ1a and HSJ1b: a role in linking the Hsp70 chaperone machine to the ubiquitin-proteasome system?

The heat-shock protein 70 chaperone machine is functionally connected to the ubiquitin-proteasome system by the co-chaperone CHIP. In this article, we discuss evidence that the neuronal DnaJ proteins HSJ1a and HSJ1b may represent a further link between the cellular protein folding and degradation machineries. We have demonstrated that HSJ1 proteins contain putative ubiquitin interaction motifs and can modulate the cellular processing of rhodopsin, a protein that is targeted for degradation by the proteasome when it is misfolded.

GnRH Neuronal Migration and Olfactory Bulb Neurite Outgrowth Are Dependent on FGF Receptor 1 Signaling, Specifically via the PI3K p110α Isoform in Chick Embryo

Fibroblast growth factor (FGF) signaling is essential for both olfactory bulb (OB) morphogenesis and the specification, migration, and maturation of the GnRH-secreting neurons. Disruption of FGF signaling contributes to Kallmann syndrome characterized by both anosmia and sexual immaturity. However, several unanswered questions remain as to which specific FGF receptor (FGFR)-1 signaling pathways are necessary for OB and GnRH neuronal development. Here, using pharmacological phosphatidylinositol 3-kinase (PI3K) isoform-specific inhibitors, we demonstrate a central role for the PI3K p110α isoform as a downstream effector of FGFR1 signaling for both GnRH neuronal migration and OB development. We show that signaling via the PI3K p110α isoform is required for GnRH neuronal migration in explant cultures of embryonic day (E) 4 chick olfactory placodes. We also show that in ovo administration of LY294002, a global PI3K inhibitor as well as an inhibitor to the PI3K p110α isoform into the olfactory placode of E3 chick embryo impairs GnRH neuronal migration toward the forebrain. In contrast, in ovo PI3K inhibitor treatment produced no obvious defects on primary olfactory sensory neuron axonal targeting and bundle formation. We also demonstrate that anosmin-1 and FGF2 induced neuronal migration of immortalized human embryonic GnRH neuroblast cells (FNC-B4-hTERT) is mediated by modulating FGFR1 signaling via the PI3K p110α isoform, specifically through phosphorylation of the PI3K downstream effectors, Akt and glycogen synthase kinase-3β. Finally, we show that neurite outgrowth and elongation of OB neurons in E10 chick OB explants are also dependent on the PI3K p110α isoform downstream of FGFR1. This study provides mechanistic insight into the etiology of Kallmann syndrome.

Refinement of the X-linked cataract locus (CXN) and gene analysis for CXN and Nance-Horan syndrome (NHS).

The X-linked congenital cataract (CXN) locus has been mapped to a 3-cM (approximately 3.5 Mb) interval on chromosome Xp22.13, which is syntenic to the mouse cataract disease locus Xcat and encompasses the recently refined Nance-Horan syndrome (NHS) locus. A positional cloning strategy has been adopted to identify the causative gene. In an attempt to refine the CXN locus, seven microsatellites were analysed within 21 individuals of a CXN family. Haplotypes were reconstructed confirming disease segregation with markers on Xp22.13. In addition, a proximal cross-over was observed between markers S3 and S4, thereby refining the CXN disease interval by approximately 400 Kb to 3.2 Mb, flanked by markers DXS9902 and S4. Two known genes (RAI2and RBBP7) and a novel gene (TL1) were screened for mutations within an affected male from the CXN family and an NHS family by direct sequencing of coding exons and intron-exon splice sites. No mutations or polymorphisms were identified, therefore excluding them as disease-causative in CXN and NHS. In conclusion, the CXN locus has been successfully refined and excludes PPEF1as a candidate gene. A further three candidates were excluded based on sequence analysis. Future positional cloning efforts will focus on the region of overlap between CXN, Xcat, and NHS.