Comparative analysis of the daily liver transcriptomes in wild nocturnal bats.

BACKGROUND: Mammals rely on the circadian clock network to regulate daily systemic metabolism and physiological activities. The liver is an important peripheral organ in mammals, and it has a unique circadian rhythm regulation process. As the only mammals that can fly, bats have attracted much research attention due to their nocturnal habits and life histories. However, few research reports exist concerning the circadian rhythms of bat liver gene expression and the relevant biological clock regulation mechanisms in the liver. RESULTS: In this study, the expression levels of liver genes of Asian particolored bats were comparatively analyzed using RNA-seq at four different time points across 24 h. A total of 996 genes were found to be rhythmic, accounting for 65% of the total number of expressed genes. The critical circadian rhythm genes Bmal1, Rev-erbα, Cry, and Ror in the liver exhibited different expression patterns throughout the day, and participated in physiological processes with rhythmic changes, including Th17 cell differentiation (ko04659), antigen processing and presentation (ko04612), the estrogen signaling pathway (ko04915), and insulin resistance (ko04931). In addition, previous studies have found that the peroxisome proliferator-activated receptor (PPAR) metabolic signaling pathway (ko03320) may play a vital role in the rhythmic regulation of the metabolic network. CONCLUSIONS: This study is the first to demonstrate diurnal changes in bat liver gene expression and related physiological processes. The results have thus further enriched our understanding of bats' biological clocks.

Comparative analysis of the daily brain transcriptomes of Asian particolored bat.

Daily rhythms are found in almost all organisms, and they comprise one of the most basic characteristics of living things. Daily rhythms are generated and mainly regulated by circadian clock. Bats have attracted interest from researchers because of their unique biological characteristics. However, little is known about the molecular underpinnings of daily rhythms in bats. In this study, we used RNA-Seq to uncover the daily rhythms of gene expression in the brains of Asian particolored bats over the 24-h day. Accordingly, four collected time points corresponding to four biological states, rest, sleep, wakefulness, and active, were selected. Several groups of genes with different expression levels in these four states were obtained suggested that different physiological processes were active at various biological states, including drug metabolism, signaling pathways, and the circadian rhythm. Furthermore, downstream analysis of all differentially expressed genes in these four states suggested that groups of genes showed daily rhythms in the bat brain. Especially for Per1, an important circadian clock gene was identified with rhythmic expression in the brain of Asian particolored bat. In summary, our study provides an overview of the brain transcriptomic differences in different physiological states over a 24-h cycle.

A secretory phospholipase D hydrolyzes phosphatidylcholine to suppress rice heading time.

Phospholipase D (PLD) hydrolyzes membrane phospholipids and is crucial in various physiological processes and transduction of different signals. Secretory phospholipases play important roles in mammals, however, whose functions in plants remain largely unknown. We previously identified a rice secretory PLD (spPLD) that harbors a signal peptide and here we reported the secretion and function of spPLD in rice heading time regulation. Subcellular localization analysis confirmed the signal peptide is indispensable for spPLD secretion into the extracellular spaces, where spPLD hydrolyzes substrates. spPLD overexpression results in delayed heading time which is dependent on its secretory character, while suppression or deficiency of spPLD led to the early heading of rice under both short-day and long-day conditions, which is consistent with that spPLD overexpression/suppression indeed led to the reduced/increased Hd3a/RFT1 (Arabidopsis Flowing Locus T homolog) activities. Interestingly, rice Hd3a and RFT1 bind to phosphatidylcholines (PCs) and a further analysis by lipidomic approach using mass spectrometry revealed the altered phospholipids profiles in shoot apical meristem, particularly the PC species, under altered spPLD expressions. These results indicate the significance of secretory spPLD and help to elucidate the regulatory network of rice heading time.

Differences in Diet and Gut Microbiota Between Lactating and Non-lactating Asian Particolored Bats (Vespertilio sinensis): Implication for a Connection Between Diet and Gut Microbiota.

In mammals, lactation is considered the most energetically costly phase for females. To meet nutritional and energy demands, lactating females usually change feeding patterns by eating food that is higher in protein and calories. Their gut microbes respond accordingly to help adapt to the changes in diet. In this study, we examined differences in diet and gut microbial composition between lactating and non-lactating Asian particolored bats (Vespertilio sinensis) using COI and 16S amplicon sequencing. When compared with non-lactating bats, we found that the diversity and composition of lactating bats' diets differed; the proportion of Diptera increased and Coleoptera and Orthoptera decreased significantly. This could be attributed to the easy availability and high protein content of Diptera. Comparative analysis of the gut microbiota of lactating and non-lactating females showed that although the diversity of gut microbiota did not change, the relative abundance of specific gut microbiota associated with a particular diet did change. For example, when the consumption of Coleoptera decreased in lactating bats, the relative abundance of Lactobacillaceae was also reduced. Lactobacillaceae are thought to be involved in the digestion of Coleopteran exoskeletons. This study suggests that during lactation, Asian particolored bats eat a diet that yields higher levels of protein, and at the same time, the abundance of specific gut microbes change to help their hosts adapt to these changes in diet.

Assessing evidence for adaptive evolution in two hearing-related genes important for high-frequency hearing in echolocating mammals.

High-frequency hearing is particularly important for echolocating bats and toothed whales. Previously, studies of the hearing-related genes Prestin, KCNQ4, and TMC1 documented that adaptive evolution of high-frequency hearing has taken place in echolocating bats and toothed whales. In this study, we present two additional candidate hearing-related genes, Shh and SK2, that may also have contributed to the evolution of echolocation in mammals. Shh is a member of the vertebrate Hedgehog gene family and is required in the specification of the mammalian cochlea. SK2 is expressed in both inner and outer hair cells, and it plays an important role in the auditory system. The coding region sequences of Shh and SK2 were obtained from a wide range of mammals with and without echolocating ability. The topologies of phylogenetic trees constructed using Shh and SK2 were different; however, multiple molecular evolutionary analyses showed that those two genes experienced different selective pressures in echolocating bats and toothed whales compared to nonecholocating mammals. In addition, several nominally significant positively selected sites were detected in the nonfunctional domain of the SK2 gene, indicating that different selective pressures were acting on different parts of the SK2 gene. This study has expanded our knowledge of the adaptive evolution of high-frequency hearing in echolocating mammals.

Phosphatidic acid (PA) binds PP2AA1 to regulate PP2A activity and PIN1 polar localization.

Phospholipase D (PLD) exerts broad biological functions in eukaryotes through regulating downstream effectors by its product, phosphatidic acid (PA). Protein kinases and phosphatases, such as mammalian target of rapamycin (mTOR), Protein Phosphatase 1 (PP1) and Protein Phosphatase 2C (PP2C), are PA-binding proteins that execute crucial regulatory functions in both animals and plants. PA participates in many signaling pathways by modulating the enzymatic activity and/or subcellular localization of bound proteins. In this study, we demonstrated that PLD-derived PA interacts with the scaffolding A1 subunit of Protein Phosphatase 2A (PP2A) and regulates PP2A-mediated PIN1 dephosphorylation in Arabidopsis. Genetic and pharmacological studies showed that both PA and PP2A participate in the regulation of auxin distribution. In addition, both the phosphorylation status and polar localization of PIN1 protein were affected by PLD inhibitors. Exogenous PA triggered the membrane accumulation of PP2AA1 and enhanced the PP2A activity at membrane, while PLD inhibition resulted in the reduced endosomal localization and perinuclear aggregation of PP2AA1. These results demonstrate the important role of PLD-derived PA in normal PP2A-mediated PIN dephosphorylation and reveal a novel mechanism, in which PA recruits PP2AA1 to the membrane system and regulates PP2A function on membrane-targeted proteins. As PA and PP2A are conserved among eukaryotes, other organisms might use similar mechanisms to mediate multiple biological processes.

Inositol polyphosphate 5-phosphatase-controlled Ins(1,4,5)P3/Ca2+ is crucial for maintaining pollen dormancy and regulating early germination of pollen.

Appropriate pollen germination is crucial for plant reproduction. Previous studies have revealed the importance of dehydration in maintaining pollen dormancy; here, we show that phosphatidylinositol pathway-controlled Ins(1,4,5)P(3)/Ca(2+) levels are crucial for maintaining pollen dormancy in Arabidopsis thaliana. An interesting phenotype, precocious pollen germination within anthers, results from a disruption of inositol polyphosphate 5-phosphatase 12 (5PT12). The knockout mutant 5pt12 has normal early pollen development and pollen dehydration, and exhibits hypersensitive ABA responses, indicating that precocious pollen germination is not caused either by abnormal dehydration or by suppressed ABA signaling. Deficiency of 5PT13 (a close paralog of 5PT12) synergistically enhances precocious pollen germination. Both basal Ins(1,4,5)P(3) levels and endogenous Ca(2+) levels are elevated in pollen from 5pt12 mutants, and 5pt12 5pt13 double mutants show an even higher precocious germination rate along with much higher levels of Ins(1,4,5)P(3)/Ca(2+). Strikingly, exogenous Ca(2+) stimulates the germination of wild-type pollen at floral stage 12, even in very low humidity, both in vitro and in vivo, and treatment with BAPTA, a [Ca(2+)](cyt) inhibitor, reduces the precocious pollen germination rates of 5pt12, 5pt13 and 5pt12 5pt13 mutants. These results indicate that the increase in the levels of Ins(1,4,5)P(3)/Ca(2+) caused by deficiency of inositol polyphosphate 5-phosphatases is sufficient to break pollen dormancy and to trigger early germination. The study reveals that independent of dehydration, the control of Ins(1,4,5)P(3)/Ca(2+) levels by Inositol polyphosphate 5-phosphatases is crucial for maintaining pollen dormancy.

Arabidopsis phosphatidylinositol monophosphate 5-kinase 2 is involved in root gravitropism through regulation of polar auxin transport by affecting the cycling of PIN proteins.

Phosphatidylinositol monophosphate 5-kinase (PIP5K) catalyzes the synthesis of PI-4,5-bisphosphate (PtdIns(4,5)P(2)) by phosphorylation of PI-4-phosphate at the 5 position of the inositol ring, and is involved in regulating multiple developmental processes and stress responses. We here report on the functional characterization of Arabidopsis PIP5K2, which is expressed during lateral root initiation and elongation, and whose expression is enhanced by exogenous auxin. The knockout mutant pip5k2 shows reduced lateral root formation, which could be recovered with exogenous auxin, and interestingly, delayed root gravity response that could not be recovered with exogenous auxin. Crossing with the DR5-GUS marker line and measurement of free IAA content confirmed the reduced auxin accumulation in pip5k2. In addition, analysis using the membrane-selective dye FM4-64 revealed the decelerated vesicle trafficking caused by PtdIns(4,5)P(2) reduction, which hence results in suppressed cycling of PIN proteins (PIN2 and 3), and delayed redistribution of PIN2 and auxin under gravistimulation in pip5k2 roots. On the contrary, PtdIns(4,5)P(2) significantly enhanced the vesicle trafficking and cycling of PIN proteins. These results demonstrate that PIP5K2 is involved in regulating lateral root formation and root gravity response, and reveal a critical role of PIP5K2/PtdIns(4,5)P(2) in root development through regulation of PIN proteins, providing direct evidence of crosstalk between the phosphatidylinositol signaling pathway and auxin response, and new insights into the control of polar auxin transport.