Genetic Informatics Laboratory / Yamazaki Group

Bacterial genome is transcribed by a single species of DNA-dependent RNA polymerase (RNAP). The genome transcription pattern is determined by controlling the utilization of a limited number of RNAP, of which the gene selectivity is modulated through two-steps of protein-protein interaction with two groups of regulatory protein, sigma factors with promoter recognition activity and transcription factors (TFs). The identification of regulatory targets for all these regulatory proteins is absolutely needed towards understanding the genome regulation at molecular level. Using the newly developed Genomic SELEX screening system, Ishihama, A. (NIG Emeritus Professor) and colleagues in Hosei University performed the systematic search of regulatory targets for all 7 sigma factors and all 300 transcriptions of the model prokaryote Escherichia coli. This is the first report on this line, describing the assembly and analysis of regulatory targets for the first group of TFs. The experiments have been carried out using a single and the same E. coli strain and using the same experimental systems in a single laboratory. To facilitate sharing these accumulated data sets with the research community, we compiled a database, “Transcription Profile of Escherichia coli” (TEC)(www.shigen.nig.ac.jp/ecoli/tec/). This pioneering research of identification of the whole set of regulatory targets for all regulatory proteins from a single organism will contribute for creation of a new paradigm in modern molecular genetics.

Using the TEC database, various types of analysis are possible:(A) search for the regulatory targets of TF. (B) genomic view of TF binding sites/intensity (C) comparison of binding sites between different TFs (D) heat map view of TF binding sites/intensity (E) Search for the consensus sequence of TF binding region.

Model Fish Genomics Resource Laboratory / Sakai Group

Molecular dissection of spermatogenesis could be facilitated by cell culture approaches that allow easy access for experimental manipulation and live imaging of specific molecules; however, technical limitations have thus far prevented the complete reconstruction of spermatogenic events in cell culture. Here, we established cell culture systems for production of functional sperm from self-renewing spermatogonial stem cells (SSCs), using zebrafish testicular hyperplasias that accumulate SSCs. By serially transplanting into immuno-deficient mutant zebrafish, we succeeded in long-term and efficient production of hyperplasias. Through improvements of culture conditions, we achieved efficient propagation of the SSCs and differentiation to sperm that gave rise to offspring. Oxygen at the concentration of air proved to be detrimental for sperm differentiation from SSCs. These results indicate that the whole spermatogenic process can be represented in cell culture, facilitating analyses of molecular mechanisms of spermatogenesis in vertebrates. This work was supported by JSPS KAKENHI (23013023, 25251034, 25114003).

Propagation and differentiation of SSCs in culture. SSCs that express green fluorescent protein grow in propagation culture (left and middle panels), while they differentiated into sperm in differentiation culture (the right panel).