Computational methods for de novo identification of gene regulation elements, such as transcription factor binding sites, have proved to be useful for deciphering genetic regulatory networks. However, despite the availability of a large number of algorithms, their strengths and weaknesses are not sufficiently understood. Here, we designed a comprehensive set of performance measures and benchmarked five modern sequence-based motif discovery algorithms using large datasets generated from Escherichia coli RegulonDB. Factors that affect the prediction accuracy, scalability and reliability are characterized. It is revealed that the nucleotide and the binding site level accuracy are very low, while the motif level accuracy is relatively high, which indicates that the algorithms can usually capture at least one correct motif in an input sequence. To exploit diverse predictions from multiple runs of one or more algorithms, a consensus ensemble algorithm has been developed, which achieved 6–45% improvement over the base algorithms by increasing both the sensitivity and specificity. Our study illustrates limitations and potentials of existing sequence-based motif discovery algorithms. Taking advantage of the revealed potentials, several promising directions for further improvements are discussed. Since the sequence-based algorithms are the baseline of most of the modern motif discovery algorithms, this paper suggests substantial improvements would be possible for them.
Nuclear importing system and nuclear factors play important roles in mediating nuclear reprogramming and zygotic gene activation. However, the components and mechanisms that mediate nuclearly specific targeting of the nuclear proteins during nuclear reprogramming and zygotic gene activation remain largely unknown. Here, we identified a novel member of the importin-␣ family, AW146299(KPNA7), which is predominantly expressed in mouse oocytes and zygotes and localizes to the nucleus or spindle. Mutation of Kpna7 gene caused reproductivity reduction and sex imbalance by inducing preferential fetal lethality in females. Parthenogenesis analysis showed that the cell cycle of activated one-cell embryos is loss of control and ahead of schedule but finally failed to develop into blastocyst stage. Further RT-PCR and epigenetic modification analysis showed that knocking out of Kpna7 induced abnormalities of gene expression (dppa2, dppa4, and piwil2) and epigenetic modifications (down-regulation of histone H3K27me3). Biochemical analysis showed that KPNA7 interacts with KPNB1 (importin-1). In summary, we identified a novel Kpna7 gene that is required for normal fertility and fecundity.
Developmental progress of germ cells through meiotic phases is closely tied to ongoing meiotic recombination. In mammals, recombination preferentially occurs in genomic regions known as hotspots; the protein that activates these hotspots is PRDM9, containing a genetically variable zinc-finger domain and a PR-SET domain with histone H3K4 trimethyltransferase activity. PRDM9 is required for fertility in mice, but little is known about its localization and developmental dynamics. Application of spermatogenic stage-specific markers demonstrates that PRDM9 accumulates in male germ-cell nuclei at pre-leptonema to early leptonema, but is no longer detectable in nuclei by late zygonema. By the pachytene stage, PRDM9-dependent histone H3K4 trimethyl marks on hotspots also disappear. PRDM9 localizes to nuclei concurrently with the deposition of meiotic cohesin complexes, but is not required for incorporation of cohesin complex proteins into chromosomal axial elements, or accumulation of normal numbers of RAD51 foci on meiotic chromatin by late zygonema. Germ cells lacking PRDM9 exhibit inefficient homology recognition and synapsis, with aberrant repair of meiotic DNA double-strand breaks and transcriptional abnormalities characteristic of meiotic silencing of unsynapsed chromatin. Together, these results on the developmental time course for nuclear localization of PRDM9 establish its direct window of function, and demonstrate the independence of chromosome axial element formation from the concurrent PRDM9-mediated activation of recombination hotspots.
The Na+/H+ antiporters (NHXs) are secondary ion transporters to exchange H+ and transfer the Na+ or K+ across membrane, they play crucial roles during plant development and stress responses. To gain insight into the functional divergence of NHX genes in poplar, eight PtNHX were identified from Populus trichocarpa genome. PtNHXs containing 10 transmembrane helices (TMH) and a hydrophilic C-terminal domain, the TMH compose a hollow cylinder to provide the channel for Na+ and H+ transport. The expression patterns and cis-acting elements showed that all the PtNHXs were response to single or multiple stresses including drought, heat, cold, salinity, MV, and ABA. Both the co-expression network and protein-protein interaction network of PtNHXs implying their functional divergence. Interestingly, although PtNHX7 and PtNHX8 were generated by whole genome duplication event, they showed significant differences in expression pattern, protein structure, co-expressed genes, and interacted proteins. Only PtNHX7 interact with CBL and CIPK, indicating PtNHX7 is the primary NHX involved in CBL-CIPK pathway during salt stress responses. Natural variation analysis based on 549 P. trichocarpa individuals indicated the frequency of SNPs in PtNHX7 was significantly higher than other PtNHXs. Our findings provide new insights into the functional divergence of NHX genes in poplar.
BackgroundIn plants, 14-3-3 proteins are encoded by a large multigene family and are involved in signaling pathways to regulate plant development and protection from stress. Although twelve Populus 14-3-3s were identified based on the Populus trichocarpa genome V1.1 in a previous study, no systematic analysis including genome organization, gene structure, duplication relationship, evolutionary analysis and expression compendium has been conducted in Populus based on the latest P. trichocarpa genome V3.0.Principal FindingsHere, a comprehensive analysis of Populus 14-3-3 family is presented. Two new 14-3-3 genes were identified based on the latest P. trichocarpa genome. In P. trichocarpa, fourteen 14-3-3 genes were grouped into ε and non-ε group. Exon-intron organizations of Populus 14-3-3s are highly conserved within the same group. Genomic organization analysis indicated that purifying selection plays a pivotal role in the retention and maintenance of Populus 14-3-3 family. Protein conformational analysis indicated that Populus 14-3-3 consists of a bundle of nine α-helices (α1-α9); the first four are essential for formation of the dimer, while α3, α5, α7, and α9 form a conserved peptide-binding groove. In addition, α1, α3, α5, α7, and α9 were evolving at a lower rate, while α2, α4, and α6 were evolving at a relatively faster rate. Microarray analyses showed that most Populus 14-3-3s are differentially expressed across tissues and upon exposure to various stresses.ConclusionsThe gene structures and their coding protein structures of Populus 14-3-3s are highly conserved among group members, suggesting that members of the same group might also have conserved functions. Microarray and qRT-PCR analyses showed that most Populus 14-3-3s were differentially expressed in various tissues and were induced by various stresses. Our investigation provided a better understanding of the complexity of the 14-3-3 gene family in poplars.
BackgroundThe continuous and non-synchronous nature of postnatal male germ-cell development has impeded stage-specific resolution of molecular events of mammalian meiotic prophase in the testis. Here the juvenile onset of spermatogenesis in mice is analyzed by combining cytological and transcriptomic data in a novel computational analysis that allows decomposition of the transcriptional programs of spermatogonia and meiotic prophase substages.ResultsGerm cells from testes of individual mice were obtained at two-day intervals from 8 to 18 days post-partum (dpp), prepared as surface-spread chromatin and immunolabeled for meiotic stage-specific protein markers (STRA8, SYCP3, phosphorylated H2AFX, and HISTH1T). Eight stages were discriminated cytologically by combinatorial antibody labeling, and RNA-seq was performed on the same samples. Independent principal component analyses of cytological and transcriptomic data yielded similar patterns for both data types, providing strong evidence for substage-specific gene expression signatures. A novel permutation-based maximum covariance analysis (PMCA) was developed to map co-expressed transcripts to one or more of the eight meiotic prophase substages, thereby linking distinct molecular programs to cytologically defined cell states. Expression of meiosis-specific genes is not substage-limited, suggesting regulation of substage transitions at other levels.ConclusionsThis integrated analysis provides a general method for resolving complex cell populations. Here it revealed not only features of meiotic substage-specific gene expression, but also a network of substage-specific transcription factors and relationships to potential target genes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2865-1) contains supplementary material, which is available to authorized users.
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