Cadmium is a common toxicant that is detrimental to many tissues. Although a number of transcriptional signatures have been revealed in different tissues after cadmium treatment, the genes involved in the cadmium caused male reproductive toxicity, and the underlying molecular mechanism remains unclear. Here we observed that the mice treated with different amount of cadmium in their rodent chow for six months exhibited reduced serum testosterone. We then performed RNA-seq to comprehensively investigate the mice testicular transcriptome to further elucidate the mechanism. Our results showed that hundreds of genes expression altered significantly in response to cadmium treatment. In particular, we found several transcriptional signatures closely related to the biological processes of regulation of hormone, gamete generation, and sexual reproduction, respectively. The expression of several testosterone synthetic key enzyme genes, such as Star, Cyp11a1, and Cyp17a1, were inhibited by the cadmium exposure. For better understanding of the cadmium-mediated transcriptional regulatory mechanism of the genes, we computationally analyzed the transcription factors binding sites and the mircoRNAs targets of the differentially expressed genes. Our findings suggest that the reproductive toxicity by cadmium exposure is implicated in multiple layers of deregulation of several biological processes and transcriptional regulation in mice. =20

Glucose is the main source of energy and carbon in organisms and plays a central role in metabolism and cellular homeostasis. However, the sensitive fluctuation of glucose in living cells is difficult to monitor. Thus, we developed a series of ratiometric, highly responsive, single fluorescent protein-based glucose sensors of wide dynamic range by combining a circularly permuted yellow fluorescent protein with a bacterial periplasmic glucose/galactose-binding protein. We used these sensors to monitor glucose transport in living Escherichia coli cells, and found that the cells take up glucose within 10 min to maintain physiological glucose levels, and observed the differences in glucose uptake and glucose metabolism between wild-type and Mlc knockout cells. These sensors can be specific and simple tools for glucose detection in vitro and non-invasive tools for real-time monitoring of glucose metabolism in vivo.

In this paper, we propose a novel and sensitive ratiometric analysis method that uses the fractional intensities of time-resolved fluorescence of genetically encoded fluorescent NADH/NAD(+) biosensors, Peredox, SoNar, and Frex. When the conformations of the biosensors change upon NADH/NAD(+) binding, the fractional intensities (alpha(i)tau(i)) have opposite changing trends. Their ratios could be exploited to quantify NADH/NAD(+) levels with a larger dynamic range and higher resolution versus commonly used fluorescence intensity and lifetime methods. Moreover, only one excitation and one emission wavelength are required for this ratiometric measurement. This eliminates problems of traditional excitation-ratiometric and emission-ratiometric methods. This method could be used to simplify the design and achieve highly sensitive analyte quantification of genetically encoded fluorescent biosensors. Wide potential applications could be developed for imaging live cell metabolism based on this new method.

High-resolution spatiotemporal imaging of histidine in single living mammalian cells faces technical challenges. Here, we developed a series of ratiometric, highly responsive, and single fluorescent protein-based histidine sensors of wide dynamic range. We used these sensors to quantify subcellular free-histidine concentrations in glucose-deprived cells and glucose-fed cells. Results showed that cytosolic free-histidine concentration was higher and more sensitive to the environment than free histidine in the mitochondria. Moreover, histidine was readily transported across the plasma membrane and mitochondrial inner membrane, which had almost similar transport rates and transport constants, and histidine transport was not influenced by cellular metabolic state. These sensors are potential tools for tracking histidine dynamics inside subcellular organelles, and they will open an avenue to explore complex histidine signaling.

Genetically encoded fluorescent sensors are widely used to visualize secondary messengers, metabolites and dynamic events in living cells. However, almost all of these sensors are based on Aequorea GFPs or GFP-like proteins, which do not correctly maturate and fluoresce under hypoxia or anoxic conditions, greatly limiting their application in biomedical research. Herein, we provide a novel strategy for design of sensors and report a series of thiol redox-sensitive sensor based on a recently discovered oxygen-independent fluorescent protein UnaG from Japanese eel. These redox sensors have large dynamic range, rapid responsiveness, a flexible "switch", and pH-independence, are particularly compatible with hypoxia conditions, and therefore represent a substantial improvement for live-cell redox measurement. We further demonstrated the versatility of these redox sensors, by simultaneously monitoring redox changes and hypoxia state in living cells, thereby proving its capability as a powerful and flexible tool for indexing multidimensional metabolism data in the context of physiological stressors and pathological states. These redox sensors are not only the first case of UnaG-based functional sensors, but also the first case of functional sensors based on non GFP-like proteins. Based on this strategy, more oxygen-independent biosensors could be developed, hence, provide new opportunities for bioimaging.

Engineered fluorescent indicators for visualizing mercury ion (Hg2+) are powerful tools to illustrate the intracellular distribution and serious toxicity of the ion. However, the sensitive and specific detection of Hg2+ in living cells and in vivo is challenging. This paper reported the development of fluorescent indicators for Hg2+ in green or red color by inserting a circularly permuted fluorescent protein into a highly mercury-specific repressor. These sensors provided a rapid, sensitive, specific, and real-time read-out of Hg2+ dynamics in solutions, bacteria, subcellular organelles of mammalian cells, and zebrafish, thereby providing a useful new method for Hg2+ detection and bioimaging. In conjunction with the hydrogen peroxide sensor HyPer, we found mercury uptake would trigger subcellular oxidative events at the single-cell level, and provided visual evidence of the causality of mercury and oxidative damage. These sensors would paint the landscape of mercury toxicity to cell functions.

Long noncoding RNAs (lncRNAs) are a large family of noncoding RNAs that play a critical role in various normal bioprocesses as well as tumorigenesis. However, the expression patterns and biological functions of lncRNAs in acute leukemia have not been well studied. Here, we performed transcriptome-wide lncRNA expression profiling of acute myeloid leukemia (AML) patient samples, along with non-leukemia control hematopoietic samples. We found that lncRNAs were differentially expressed in AML samples relative to control samples. Notably, we identified that lncRNAs upregulated in AML (relative to the control samples) are associated with a lower degree of DNA methylation and a higher ratio of being bound by transcription factors such as SP1, STAT4, ATF-2 and ELK-1 compared with those downregulated in AML. Moreover, an enrichment of H3K4me3 and a depletion of H3K27me3 were observed in upregulated lncRNAs in AML. Expression patterns of three types of lncRNAs (antisense, enhancer and intergenic lncRNAs) have previously been characterized. Of the identified lncRNAs, we found that high expression level lncRNA LOC285758 is associated with the poor prognosis in AML patients. Furthermore, we found that LOC285758 regulates proliferation of AML cell lines by enhancing the expression of HDAC2, a key factor in carcinogenesis. Collectively, our study depicts a landscape of important lncRNAs in AML and provides novel potential therapeutic targets and prognostic markers for AML treatment.=20

Breast cancer has been classified into several intrinsic molecular subtypes on the basis of genetic and epigenetic factors. However, knowledge about histone modifications that contribute to the classification and development of biologically distinct breast cancer subtypes remains limited. Here we compared the genome-wide binding patterns of H3K4me3 and H3K27me3 between human mammary epithelial cells and three breast cancer cell lines representing the luminal, HER2, and basal subtypes. We characterized thousands of unique binding events as well as bivalent chromatin signatures unique to each cancer subtype, which were involved in different epigenetic regulation programs and signaling pathways in breast cancer progression. Genes linked to the unique histone mark features exhibited subtype-specific expression patterns, both in cancer cell lines and primary tumors, some of which were confirmed by qPCR in our primary cancer samples. Finally, histone mark-based gene classifiers were significantly correlated with relapse-free survival outcomes in patients. In summary, we have provided a valuable resource for the identification of novel biomarkers of subtype classification and clinical prognosis evaluation in breast cancers.