Project description:Monobromobimane (mBBr) labelling: mBBr labelling was carried out either immediately prior to 42°C, two hour heat shock, or immediately following heat shock. Medium was removed from culture flasks and cells were washed using PBS. Cells were then labelled by incubation at 37°C with 6 mL of 400 µM mBBr (Sigma-Aldrich, #B4380) for 10 minutes. Following labeling, 6 ml of 2 mM L-glutathione reduced (GSH, SigmaAldrich, #G4251) in PBS was added to quench the mBBr reaction. The quenched mBBr solution was removed and cells were washed with PBS. Mass spectrometry (MS) sample preparation, analysis and data processing: Six 1.6 mm steel beads (Next Advance Inc.) were added to the cell pellet tube with 30 µL SL-DOC (1.1% (w/w) sodium dodecyl sulfate (Sigma), 0.3% (w/w) sodium deoxycholate (Sigma), 25 mM ammonium bicarbonate (AB, Fluka), in de-ionised (DI) water containing 0.1% (v/v) protease inhibitor cocktail (Sigma), and 0.1% (v/v) phosphotase inhibitor cocktail (Sigma). Cells were homogen
Project description:In the fire ant Solenopsis invicta, a colony queen number is determined by the founding queen's genotypes at the 13 Mb supergene with the non-recombining variants SB and Sb. Single-queen colonies are always headed by SB/SB queens while multiple-queens colonies are always headed by SB/Sb queens. The two variants of the supergene, SB and Sb are completely linked to the two alleles (B and b) of the gene Gp-9. SB/SB and SB/Sb queens differ in many physiological traits including their maturation rate and odor. To explain why SB/SB and SB/Sb queens have different odors, and why SB/SB virgins mature faster and accumulate more fat, we measured expression of ~6000 genes in virgin queens 1 and 11 days after eclosion and in reproductive queens. Keywords: fire ants, Solenopsis invicta, Supergene, queen, Gp-9, social form, maturation, fat storage, queen odor, cuticular hydrocarbon, worker discrimination, monogyne, polygyne, transposon, chemical signaling
Project description:Queen discrimination behavior in the red imported fire ant Solenopsis invicta maintains its two types of societies: colonies with one (monogyne) or many (polygyne) queens, yet the underlying genetic mechanism is poorly understood. This behavior is controlled by two supergene alleles, SB and Sb, with ~600 genes. Polygyne workers, having either the SB/SB or SB/Sb genotype, accept additional SB/Sb queens into their colonies but kill SB/SB queens. While monogyne workers, all SB/SB, reject all additional queens regardless of genotype. Because the SB and Sb alleles do not recombine, it is difficult to determine which genes within the supergene mediate this differential worker behavior. We hypothesized that the alternate worker genotypes sense queens differently because of different patterns of gene expression in their main sensory organ, the antennae. To identify such differentially expressed genes, we sequenced RNA from four biological replicates of pooled antennae from three groups of workers: monogyne SB/SB, polygyne SB/SB, and polygyne SB/Sb. We identified 81 differentially expressed protein coding genes with 14 encoding potential odor metabolism and perception proteins. We focused on the two differentially expressed odorant perception genes: an odorant binding protein SiOBP12 and an odorant receptor SiOR463. We found that the SiOR463 was lost in the Sb-genome. In contrast, the SiOBP12 has an Sb-specific duplication SiOBP12b’, which was expressed in the SB/Sb worker antennae, while both paralogs SiOBP12 and SiOBP12b’ were expressed in the body. This result indicates that SiOBP12b’ has gained an antennal promoter or enhancer and suggests neofunctionalization, perhaps for queen discrimination behavior.
Project description:Queen discrimination behavior in the red imported fire ant Solenopsis invicta maintains its two types of societies: colonies with one (monogyne) or many (polygyne) queens, yet the underlying genetic mechanism is poorly understood. This behavior is controlled by two supergene alleles, SB and Sb, with ~600 genes. Polygyne workers, having either the SB/SB or SB/Sb genotype, accept additional SB/Sb queens into their colonies but kill SB/SB queens. While monogyne workers, all SB/SB, reject all additional queens regardless of genotype. Because the SB and Sb alleles do not recombine, it is difficult to determine which genes within the supergene mediate this differential worker behavior. We hypothesized that the alternate worker genotypes sense queens differently because of different patterns of gene expression in their main sensory organ, the antennae. To identify such differentially expressed genes, we sequenced RNA from four biological replicates of pooled antennae from three groups of workers: monogyne SB/SB, polygyne SB/SB, and polygyne SB/Sb. We identified 81 differentially expressed protein coding genes with 14 encoding potential odor metabolism and perception proteins. We focused on the two differentially expressed odorant perception genes: an odorant binding protein SiOBP12 and an odorant receptor SiOR463. We found that the SiOR463 was lost in the Sb-genome. In contrast, the SiOBP12 has an Sb-specific duplication SiOBP12b’, which was expressed in the SB/Sb worker antennae, while both paralogs SiOBP12 and SiOBP12b’ were expressed in the body. This result indicates that SiOBP12b’ has gained an antennal promoter or enhancer and suggests neofunctionalization, perhaps for queen discrimination behavior.
Project description:Queen discrimination behavior in the red imported fire ant Solenopsis invicta maintains its two types of societies: colonies with one (monogyne) or many (polygyne) queens, yet the underlying genetic mechanism is poorly understood. This behavior is controlled by two supergene alleles, SB and Sb, with ~600 genes. Polygyne workers, having either the SB/SB or SB/Sb genotype, accept additional SB/Sb queens into their colonies but kill SB/SB queens. While monogyne workers, all SB/SB, reject all additional queens regardless of genotype. Because the SB and Sb alleles do not recombine, it is difficult to determine which genes within the supergene mediate this differential worker behavior. We hypothesized that the alternate worker genotypes sense queens differently because of different patterns of gene expression in their main sensory organ, the antennae. To identify such differentially expressed genes, we sequenced RNA from four biological replicates of pooled antennae from three groups of workers: monogyne SB/SB, polygyne SB/SB, and polygyne SB/Sb. We identified 81 differentially expressed protein coding genes with 14 encoding potential odor metabolism and perception proteins. We focused on the two differentially expressed odorant perception genes: an odorant binding protein SiOBP12 and an odorant receptor SiOR463. We found that the SiOR463 was lost in the Sb-genome. In contrast, the SiOBP12 has an Sb-specific duplication SiOBP12b’, which was expressed in the SB/Sb worker antennae, while both paralogs SiOBP12 and SiOBP12b’ were expressed in the body. This result indicates that SiOBP12b’ has gained an antennal promoter or enhancer and suggests neofunctionalization, perhaps for queen discrimination behavior.
Project description:Sleep bruxism (SB) is a disorder in which patients repeatedly make involuntary jaw movements during sleep. In this study, we performed bulk RNA sequencing (CTRL:SB = 3:3) using iPS cell-derived neurons established from SB patients to investigate gene expression patterns specific to SB patients.
Project description:Sleep bruxism (SB) is a disorder in which patients repeatedly make involuntary jaw movements during sleep. In this study, we performed single nucleus RNA sequencing (CTRL:SB = 1:1) using iPS cell-derived neurons established from SB patients to investigate gene expression of each neural subtype specific to SB patients.