Ashiq Hussain

Positive Darwinian selection and the birth of an olfactory receptor clade in teleosts

Trace amine-associated receptors (TAARs) in mammals recently have been shown to function as olfactory receptors. We have delineated the taar gene family in jawless, cartilaginous, and bony fish (zero, 2, and >100 genes, respectively). We conclude that taar genes are evolutionary much younger than the related OR and ORA/V1R olfactory receptor families, which are present already in lamprey, a jawless vertebrate. The 2 cartilaginous fish genes appear to be ancestral for 2 taar classes, each with mammalian and bony fish (teleost) representatives. Unexpectedly, a whole new clade, class III, of taar genes originated even later, within the teleost lineage. Taar genes from all 3 classes are expressed in subsets of zebrafish olfactory receptor neurons, supporting their function as olfactory receptors. The highly conserved TAAR1 (shark, mammalian, and teleost orthologs) is not expressed in the olfactory epithelium and may constitute the sole remnant of a primordial, nonolfactory function of this family. Class III comprises three-fourths of all teleost taar genes and is characterized by the complete loss of the aminergic ligand-binding motif, stringently conserved in the other 2 classes. Two independent intron gains in class III taar genes represent extraordinary evolutionary dynamics, considering the virtual absence of intron gains during vertebrate evolution. The dN/dS analysis suggests both minimal global negative selection and an unparalleled degree of local positive selection as another hallmark of class III genes. The accelerated evolution of class III teleost taar genes conceivably might mark the birth of another olfactory receptor gene family.

High-affinity olfactory receptor for the death-associated odor cadaverine

Significance Cadaverine and putrescine, two diamines emanating from decaying flesh, are strongly repulsive odors to humans but serve as innate attractive or social cues in other species. Here we show that zebrafish, a vertebrate model system, exhibit powerful and innate avoidance behavior to both diamines, and identify a high-affinity olfactory receptor for cadaverine. Carrion smell is strongly repugnant to humans and triggers distinct innate behaviors in many other species. This smell is mainly carried by two small aliphatic diamines, putrescine and cadaverine, which are generated by bacterial decarboxylation of the basic amino acids ornithine and lysine. Depending on the species, these diamines may also serve as feeding attractants, oviposition attractants, or social cues. Behavioral responses to diamines have not been investigated in zebrafish, a powerful model system for studying vertebrate olfaction. Furthermore, olfactory receptors that detect cadaverine and putrescine have not been identified in any species so far. Here, we show robust olfactory-mediated avoidance behavior of zebrafish to cadaverine and related diamines, and concomitant activation of sparse olfactory sensory neurons by these diamines. The large majority of neurons activated by low concentrations of cadaverine expresses a particular olfactory receptor, trace amine-associated receptor 13c (TAAR13c). Structure-activity analysis indicates TAAR13c to be a general diamine sensor, with pronounced selectivity for odd chains of medium length. This receptor can also be activated by decaying fish extracts, a physiologically relevant source of diamines. The identification of a sensitive zebrafish olfactory receptor for these diamines provides a molecular basis for studying neural circuits connecting sensation, perception, and innate behavior.

Ionotropic Chemosensory Receptors Mediate the Taste and Smell of Polyamines

The ability to find and consume nutrient-rich diets for successful reproduction and survival is fundamental to animal life. Among the nutrients important for all animals are polyamines, a class of pungent smelling compounds required in numerous cellular and organismic processes. Polyamine deficiency or excess has detrimental effects on health, cognitive function, reproduction, and lifespan. Here, we show that a diet high in polyamine is beneficial and increases reproductive success of flies, and we unravel the sensory mechanisms that attract Drosophila to polyamine-rich food and egg-laying substrates. Using a combination of behavioral genetics and in vivo calcium imaging, we demonstrate that Drosophila uses multisensory detection to find and evaluate polyamines present in overripe and fermenting fruit, their favored feeding and egg-laying substrate. In the olfactory system, two coexpressed ionotropic receptors (IRs), IR76b and IR41a, mediate the long-range attraction to the odor. In the gustatory system, multimodal taste sensation by IR76b receptor and GR66a bitter receptor neurons is used to evaluate quality and valence of the polyamine providing a mechanism for the fly’s high attraction to polyamine-rich and sweet decaying fruit. Given their universal and highly conserved biological roles, we propose that the ability to evaluate food for polyamine content may impact health and reproductive success also of other animals including humans.

Neuropeptides Modulate Female Chemosensory Processing upon Mating in Drosophila.

A female’s reproductive state influences her perception of odors and tastes along with her changed behavioral state and physiological needs. The mechanism that modulates chemosensory processing, however, remains largely elusive. Using Drosophila, we have identified a behavioral, neuronal, and genetic mechanism that adapts the senses of smell and taste, the major modalities for food quality perception, to the physiological needs of a gravid female. Pungent smelling polyamines, such as putrescine and spermidine, are essential for cell proliferation, reproduction, and embryonic development in all animals. A polyamine-rich diet increases reproductive success in many species, including flies. Using a combination of behavioral analysis and in vivo physiology, we show that polyamine attraction is modulated in gravid females through a G-protein coupled receptor, the sex peptide receptor (SPR), and its neuropeptide ligands, MIPs (myoinhibitory peptides), which act directly in the polyamine-detecting olfactory and taste neurons. This modulation is triggered by an increase of SPR expression in chemosensory neurons, which is sufficient to convert virgin to mated female olfactory choice behavior. Together, our data show that neuropeptide-mediated modulation of peripheral chemosensory neurons increases a gravid female’s preference for important nutrients, thereby ensuring optimal conditions for her growing progeny.

Elimination of a ligand gating site generates a supersensitive olfactory receptor

Olfaction poses one of the most complex ligand-receptor matching problems in biology due to the unparalleled multitude of odor molecules facing a large number of cognate olfactory receptors. We have recently deorphanized an olfactory receptor, TAAR13c, as a specific receptor for the death-associated odor cadaverine. Here we have modeled the cadaverine/TAAR13c interaction, exchanged predicted binding residues by site-directed mutagenesis, and measured the activity of the mutant receptors. Unexpectedly we observed a binding site for cadaverine at the external surface of the receptor, in addition to an internal binding site, whose mutation resulted in complete loss of activity. In stark contrast, elimination of the external binding site generated supersensitive receptors. Modeling suggests this site to act as a gate, limiting access of the ligand to the internal binding site and thereby downregulating the affinity of the native receptor. This constitutes a novel mechanism to fine-tune physiological sensitivity to socially relevant odors.

Inhibition of oxidative stress in cholinergic projection neurons fully rescues aging-associated olfactory circuit degeneration in Drosophila

Loss of the sense of smell is among the first signs of natural aging and neurodegenerative diseases such as Alzheimer’s and Parkinson’s. Cellular and molecular mechanisms promoting this smell loss are not understood. Here, we show that Drosophila melanogaster also loses olfaction before vision with age. Within the olfactory circuit, cholinergic projection neurons show a reduced odor response accompanied by a defect in axonal integrity and reduction in synaptic marker proteins. Using behavioral functional screening, we pinpoint that expression of the mitochondrial reactive oxygen scavenger SOD2 in cholinergic projection neurons is necessary and sufficient to prevent smell degeneration in aging flies. Together, our data suggest that oxidative stress induced axonal degeneration in a single class of neurons drives the functional decline of an entire neural network and the behavior it controls. Given the important role of the cholinergic system in neurodegeneration, the fly olfactory system could be a useful model for the identification of drug targets.

Correction: Ionotropic Chemosensory Receptors Mediate the Taste and Smell of Polyamines

[This corrects the article DOI: 10.1371/journal.pbio.1002454.].

Computational modeling and functional characterization of a GgChi: A class III chitinase from corms of Gladiolus grandiflorus

The present study describes the predicted model and functional characterization of an endochitinase (30 kDa) from corms of Gladiolus grandiflorus. ESI‐QTOF‐MS generated peptide showed 96% sequence homology with family 18, Class III acidic endochitinase of Gladiolus gandavensis. Purified G. grandiflorus chitinase (GgChi) hydrolyzed 4‐methylumbelliferyl β‐d‐N,N′,N′′‐triacetylchitotriose substrate showing specific endochitinase activity. Since no structural details of GgChi were available in the Protein Data Bank (PDB), a homology model was predicted using the coordinate information of Crocus vernus chitinase (PDB ID: 3SIM). Ramachandran plot indicated 84.5% in most favored region, 14.8% in additional and 0.6% in generously allowed region while no residue in disallowed region. The predicted structure indicated a highly conserved (β/α)8 (TIM barrel) structure similar to the family 18, class III chitinases. The GgChi also showed sequence and structural homologies with other active chitinases. The GgChi (50 μg/disc) showed no antibacterial activity, but did provide mild growth inhibition of phytopathogenic fungus Fusarium oxysporum at a concentration of 500 μg/well Similarly, insect toxicity bioassays of GgChi (50 μg) against nymphs of Bemisia tabaci showed 14% reduction in adult emergence and 14% increase in mortality rate in comparison to control values. The GgChi (1.5 mg) protein showed significant reduction in a population of flour beetle (Tribolium castaneum) after 35 days, but lower reactivity against rice weevil (Sitophilus oryzae). The results of this study provide detai.led insight on functional characterization of a family 18 class III acidic plant endochitinase.

MOLECULAR MODEL OF CYTOTOXIN-1 FROM NAJA MOSSAMBICA MOSSAMBICA VENOM IN COMPLEX WITH CHYMOTRYPSIN.

Snake venom is a myriad of biologically active proteins and peptides. Three finger toxins are highly conserved in their molecular structure, but interestingly possess diverse biological functions. During the course of evolution the introduction of subtle mutations in loop regions and slight variations in the three dimensional structure, has resulted in their functional versatility. Cytotoxin-1 (UniProt ID: P01467), isolated from Naja mossambica mossambica, showed the potential to inhibit chymotrypsin and the chymotryptic activity of the 20S proteasome. In the present work we describe a molecular model of cytotoxin-1 in complex with chymotrypsin, prepared by the online server ClusPro. Analysis of the molecular model shows that Cytotoxin-1 (P01467) binds to chymotrypsin through its loop I located near the N-terminus. The concave side of loop I of the toxin fits well in the substrate binding pocket of the protease. We propose Phe10 as the dedicated P1 site of the ligand. Being a potent inhibitor of the 20S proteasome, cytotoxin-1 (P01467) can serve as a potential antitumor agent. Already snake venom cytotoxins have been investigated for their ability as an anticancer agent. The molecular model of cytotoxin-1 in complex with chymotrypsin provides important information towards understanding the complex formation.