Identification of pheromone-carrying protein in the preorbital gland post in the endangered Indian male Blackbuck Antelope cervicapra L.

In mammals, a low molecular mass protein (17-20 KDa) reported from the pheromone sources such as urine, saliva, glandular secretion, etc., as ligand-carrier (pheromone carrier) has been associated with chemo-communication. Since the preorbital gland post is one of the major pheromone sources in Indian Blackbuck, an endangered species, we assumed thta it possibly contains low molecular mass protein for chemical communication. Hence, we investigated the preorbital gland post in territorial and non-territorial male blackbucks for such low molecular mass proteins adopting SDS-PAGE and LC-MS/MS analysis. The total content of protein was higher in the post of territorial males than non-territorial males of adult and sub-adult. In fact, the protein profiles such as 17, 21, 25, 42 and 61 kDa were noted in the gland secretion of territorial and non-territorial males. The intensity of the 17 kDa protein band was higher in territorial males than non-territorial males. In-gel trypsin digestion of the 17 kDa band was processed and subjected to LC-MS/MS and SEQUEST analyses. The results of LC-MS/MS and SEQUEST search showed the presence of α2u-globulin in the 17 kDa band. In addition, the identified α2u-globulin sequence possessed GDW residues, which are the characteristic signature for lipocalin family. Since the α2u-globulin has been reported from the pheromone-carrying proteins in some mammals, this protein may carry the volatiles (pheromone compounds) in male Blackbucks preorbital gland to evoke the scent marking for maintaining territoriality (home range) and attraction towards female, through the secretion of glandular protein.

Identification of glandular (preputial and clitoral) proteins in house rat (Rattus rattus) involved in pheromonal communication.

Proteins (18-20 kDa) belonging to lipocalin family have been reported to act as carriers for ligands binding to pheromones in mouse urine, pig saliva, hamster vaginal fluid and human sweat, that are involved in pheromonal communication. As the preputial gland is a major pheromonal source, the present study was aimed to detect the specific protein bands (around 18-20 kDa) in the preputial and clitoral glands of the house rat, R. rattus. The amount of protein was higher in preputial gland of the male than that of female (clitoral) gland. A 20 kDa protein was noted in male and female glands; however, the intensity of the band was much higher in male than in female. In addition, 70, 60, 35 kDa bands, identified in male preputial gland, were absent in females. The presence of higher concentration of glandular proteins in the male preputial gland suggests that male rats may depend more on these glandular proteins for the maintenance of reproductive and dominance behaviours. The results further suggest that these glandular proteins (20 kDa) may act as a carrier for ligand binding.

Characterization of urinary volatiles in Swiss male mice (Mus musculus): bioassay of identified compounds.

The present study was carried out to investigate the chemical nature of the urine of male mice and to assess its bioactivity. Urine of mature male mice was extracted with dichloromethane (1 : 1 ratio v/v) and analysed by gas-chromatography linked mass-spectrometry (GC-MS). Ten different compounds such as alkanes, alcohols, etc. were detected in the urine. Among the ten, five compounds are specific to males namely, 3-cyclohexene-1-methanol (I), 3-amino-s-triazole (II), 4-ethyl phenol (III), 3-ethyl-2,7-dimethyl octane (IV) and 1-iodoundecane (V). The compound, 4-ethylphenol, has been previously reported in several strains of male mice. Furthermore, the compounds (II) and (IV) are closely similar to 2-sec-butylthiazole and dehydro-exo-brevicomin compounds which have already been reported in male mice. Bioassay revealed that compounds (II), (III) and (IV) were responsible for attracting females and in inducing aggression towards males, as compared to the other compounds i.e. (I) and (V). The results indicate that these three volatiles (II, III and IV) of male mice appear to act as attractant of the opposite sex.

Urinary proteins and pheromonal communication in mammals.

Urinary proteins play a significant role as pheromones and pheromone-binders in mammalian reproduction and social behaviour. The present study was carried out to quantify the urinary proteins in five different mammalian species viz mouse, rat, rabbit, bovine and human. The results revealed that the male rodents excrete large amounts of urinary protein as compared to that of other mammals. In addition, the male mammals excrete a higher quantity of protein than do the females., suggesting the role of androgens in excretion of protein. The presence of higher concentration of urinary proteins in rodents suggests that the rodents depend more on urinary proteins for olfactory/social communication.