Single-cell sequencing technologies in bladder cancer research: Applications and challenges.

Bladder cancer is among the most common malignant tumors with highly heterogeneous molecular characteristics. Despite advancements of the available therapeutic options, several bladder cancer patients exhibit unsatisfactory clinical outcomes. The lack of specific biomarkers for effective targeted therapy or immunotherapy remains a major obstacle in treating bladder cancer. The rapid development of single-cell techniques is transforming our understanding of the intra-tumoral heterogeneity, thereby providing us with a powerful high-throughput sequencing tool that can reveal tumorigenesis, progression, and invasion in bladder tumors. In this review, we summarise and discuss how single-cell sequencing technologies have been applied in bladder cancer research, to advance our collective knowledge on the heterogeneity of bladder tumor cells, as well as to provide new insights into the complex ecosystem of the tumor microenvironment. The application of single-cell approaches also uncovers the therapeutic resistance mechanism in bladder cancer and facilitates the detection of urinary-exfoliated tumor cells. Moreover, benefiting from the powerful technical advantages of single-cell techniques, several key therapeutic targets and prognostic models of bladder cancer have been identified. It is hoped that this paper can provide novel insights into the precision medicine of bladder cancer.

Single-cell RNA-seq of UVB-radiated skin reveals landscape of photoaging-related inflammation and protection by vitamin D.

Ultraviolet rays are a potential threat to nature. It can accelerate skin aging by causing skin damage, cell infiltration, and inflammation. The present study investigated UV-irradiated mouse skin through single-cell sequencing. We observed that UV-irradiated mouse skin mainly induced inflammation of fibroblasts and demonstrated differential gene expression. Cell prediction revealed the significance of macrophages in tissue repair. Furthermore, cell culture studies substantiated vitamin D-induced inhibitory effect on skin inflammation. These findings thus indicate some references for skin photo-protection.

Complex Molecular Evolution and Expression of Expansin Gene Families in Three Basic Diploid Species of Brassica.

Expansins are a kind of structural proteins of the plant cell wall, and they enlarge cells by loosening the cell walls. Therefore, expansins are involved in many growth and development processes. The complete genomic sequences of Brassica rapa, Brassica oleracea and Brassica nigra provide effective platforms for researchers to study expansin genes, and can be compared with analogues in Arabidopsis thaliana. This study identified and characterized expansin families in B. rapa, B. oleracea, and B. nigra. Through the comparative analysis of phylogeny, gene structure, and physicochemical properties, the expansin families were divided into four subfamilies, and then their expansion patterns and evolution details were explored accordingly. Results showed that after the three species underwent independent evolution following their separation from A. thaliana, the expansin families in the three species had increased similarities but fewer divergences. By searching divergences of promoters and coding sequences, significant positive correlations were revealed among orthologs in A. thaliana and the three basic species. Subsequently, differential expressions indicated extensive functional divergences in the expansin families of the three species, especially in reproductive development. Hence, these results support the molecular evolution of basic Brassica species, potential functions of these genes, and genetic improvement of related crops.

Effects of intrauterine exposure to 2,3',4,4',5-pentachlorobiphenyl on the reproductive system and sperm epigenetic imprinting of male offspring.

Polychlorinated biphenyls (PCBs) are a class of persistent organic environmental pollutants with a total of 209 homologs. The homolog 2,3',4,4',5-pentachlorobiphenyl (PCB118) is one of the most important dioxin-like PCBs and is highly toxic. PCB118 can accumulate in human tissues, serum and breast milk, which leads to direct exposure of the fetus during development. In the present study, pregnant mice were exposed to 0, 20 and 100 µg/kg/day of PCB118 during the stage of fetal primordial germ cell migration. Compared with the control group, we found morphological alterations of the seminiferous tubules and a higher sperm deformity rate in the male offspring in the treatment groups. Furthermore, the methylation patterns in the treatment groups of the imprinted genes H19 and Gtl2 in the sperm were altered in the male offspring. We also characterized the disturbance of the expression levels of DNA methyltransferase 1 (Dnmt1), Dnmt3a, Dnmt3b, Dnmt3l, and Uhrf1. The results indicated that intrauterine exposure to low doses of PCB118 could significantly damage the reproductive health of the male offspring. Therefore, attention should be paid to the adverse effects of PCB118 exposure during pregnancy on the reproductive system of male offspring.

Molecular characterization and expression analysis reveal the roles of Cys2/His2 zinc-finger transcription factors during flower development of Brassica rapa subsp. chinensis.

KEY MESSAGE: Conserved motif, gene structure, expression and interaction analysis of C2H2-ZFPs in Brassica rapa, and identified types of genes may play essential roles in flower development, and BrZFP38 was proved to function in flower development by affecting pollen formation. Flower development plays a central role in determining the reproduction of higher plants, and Cys2/His2 zinc-finger proteins (C2H2-ZFPs) widely participate in the transcriptional regulation of flower development. C2H2-ZFPs with various structures are the most widespread DNA-binding transcription factors in plants. In this study, conserved protein motif and gene structures were analyzed to investigate systematically the molecular features of Brassica rapa C2H2-ZFP genes. Expression of B. rapa C2H2-ZFPs in multiple tissues showed that more than half of the family members with different types ZFs were expressed in flowers. The specific expression profiles of these C2H2-ZFPs in different B. rapa floral bud stages were further evaluated to identify their potential roles in flower development. Interaction networks were constructed in B. rapa based on the orthology of flower-related C2H2-ZFP genes in Arabidopsis. The putative cis-regulatory elements in the promoter regions of these C2H2-ZFP genes were thoroughly analyzed to elucidate their transcriptional regulation. Results showed that the orthologs of known-function flower-related C2H2-ZFP genes were conserved and differentiated in B. rapa. A C2H2-ZFP was proved to function in B. rapa flower development. Our study provides a systematic investigation of the molecular characteristics and expression profiles of C2H2-ZFPs in B. rapa and promotes further work in function and transcriptional regulation of flower development.

Arabidopsis Cys2/His2 zinc-finger protein MAZ1 is essential for intine formation and exine pattern.

Cys2/His2 zinc-finger protein (C2H2-ZFP) is widely involved in the reproductive development of plants, but its role in pollen development is still elusive. Here, we identified a pollen-related C2H2-ZFP gene named as MALE FERTILITY-ASSOCIATED ZINC FINGER PROTEIN 1 (MAZ1), which was first isolated from Arabidopsis thaliana. MAZ1 showed a preferential expression pattern in early anther development. Its mutation resulted in aberrant primexine deposition at the tetrad stage, followed by a defective multiple-layer pattern of exine with irregular baculum and no tectum. Furthermore, microspore development was arrested, and no intine layer was formed. These developmental defects led to fertility reduction and pollen abortion. This study reveals the essential role of MAZ1 in pollen wall development.

3,3',4,4',5-Pentachlorobiphenyl influences mitochondrial apoptosis pathway in granulosa cells.

3,3',4,4',5-Polychlorinated biphenyl (PCB126) is a persistent organic environmental pollutant which can affect various biological activities of organisms, such as immunity, neurological function, and reproduction. In our study, we aimed to investigate the effects of PCB126 on granulosa cells (GCs). GCs were collected from ovaries in PMSG-treated mice, after 24 hours culture. GCs were then incubated with 10 pg/mL, 100 pg/mL, and 10 ng/mL of PCB126 for another 24 hours. Following these steps, exposed GCs were collected for further experimentation. Our data showed that the number of GCs in the 10 ng/mL PCB126 decreased. Meanwhile, pyknotic nuclei and condensed chromatin increased, while the apoptotic cells in the 10 ng/mL PCB126 group were significantly increased. Furthermore, the expression of the apoptotic executive protein caspase-3 increased after PCB126 treatment. The expression of Bax, Bcl-2, and Bim related to the mitochondrial apoptosis pathway were also influenced to different degrees. Thus, our data suggested that PCB126 affect the GCs apoptosis, and mitochondrial apoptosis pathway was involved in this process.

Cys2/His2 Zinc-Finger Proteins in Transcriptional Regulation of Flower Development.

Flower development is the core of higher-plant ontogenesis and is controlled by complex gene regulatory networks. Cys2/His2 zinc-finger proteins (C2H2-ZFPs) constitute one of the largest transcription factor families and are highly involved in transcriptional regulation of flowering induction, floral organ morphogenesis, and pollen and pistil maturation. Nevertheless, the molecular mechanism of C2H2-ZFPs has been gradually revealed only in recent years. During flowering induction, C2H2-ZFPs can modify the chromatin of FLOWERING LOCUS C, thereby providing additional insights into the quantification of transcriptional regulation caused by chromatin regulation. C2H2-ZFPs are involved in cell division and proliferation in floral organ development and are associated with hormonal regulation, thereby revealing how a flower is partitioned into four developmentally distinct whorls. The studies reviewed in this work integrate the information from the endogenous, hormonal, and environmental regulation of flower development. The structure of C2H2-ZFPs determines their function as transcriptional regulators. The findings indicate that C2H2-ZFPs play a crucial role in flower development. In this review, we summarize the current understanding of the structure, expression, and function of C2H2-ZFPs and discuss their molecular mechanism in flower development.

Over-expression of miR158 causes pollen abortion in Brassica campestris ssp. chinensis.

KEY MESSAGE: We identified and cloned the two precursors of miR158 and its target gene in Brassica campestris ssp. chinensis, which both had high relative expression in the inflorescences. Further study revealed that over-expression of miR158 caused reduced pollen varbility, which was caused by the degradation of pollen contents from the binucleate microspore stage. These results first suggest the role of miR158 in pollen development of Brassica campestris ssp. chinensis. MicroRNAs (miRNAs) play crucial roles in many important growth and development processes both in plants and animals by regulating the expression of their target genes via mRNA cleavage or translational repression. In this study, miR158, a Brassicaceae specific miRNA, was functionally characterized with regard to its role in pollen development of non-heading Chinese cabbage (Brassica campestris ssp. chinensis). Two family members of miR158 in B. campestris, namely bra-miR158a1 and bra-miR158a2, and their target gene bra027656, which encodes a pentatricopeptide repeat (PPR) containing protein, were identified. Then, qRT-PCR analysis and GUS-reporter system revealed that both bra-miR158 and its target gene had relatively high expression levels in the inflorescences. Further study revealed that over-expression of miR158 caused reduced pollen varbility and pollen germination ratio, and the degradation of pollen contents from the binucleate microspore stage was also found in those deformed pollen grains, which led to pollen shrinking and collapse in later pollen development stage. These results first shed light on the importance of miR158 in pollen development of Brassica campestris ssp. chinensis.