bioRxiv ScienceSearch

Biology subjects

Trimmer, K. A.

Publications and source records attributed to Trimmer, K. A..

2 recordsLinked to original sources

Drosha Regulates Oogenesis and microRNAs Germline Autonomously and Non autonomously in C. elegans

Small non-coding RNAs regulate multiple aspects of development including germ cell development. The microRNA pathway genes Dicer, Drosha and Pasha have been shown to regulate oocyte meiotic maturation in C. elegans. However, Dicer controls oocyte meiotic maturation through endo-siRNAs, rather than microRNAs. A repertoire of Drosha-dependent oocyte-expressed microRNAs were identified which regulate various aspects of oogenesis but not oocyte meiotic maturation. These data lead to the following models: (a) microRNAs function redundantly to regulate oocyte meiotic maturation, (b) Drosha and microRNAs function germline non-autonomously to regulate meiotic maturation. We investigated these models and observed that Drosha regulates oocyte meiotic maturation in a germline non-autonomous manner. Additionally, we uncovered a role for Drosha in regulating pachytene progression and oocyte development in a germline autonomous manner through miR-35 family and miR-51 respectively. Interestingly we also find that though Drosha-dependent oocyte-expressed microRNAs, miR-61 and miR-72, are sufficient to regulate pachytene progression and oocyte development respectively, they are generated in a germline non-autonomous manner. Collectively these data reveal a Drosha-dependent microRNA circuit, which coordinates oocyte development germline autonomously as well as through soma-germline communication.

developmental biology

CreLite: An Optogenetically Controlled Cre/loxP System Using Red Light

Precise manipulation of gene expression with temporal and spatial control is essential for functional studies and the determination of cell lineage relationships in complex biological systems. The Cre-loxP system is commonly used for gene manipulation at desired times and places. However, specificity is dependent on the availability of tissue- or cell-specific regulatory elements used in combination with Cre or CreER (tamoxifen-inducible). Here we present CreLite, an optogenetically-controlled Cre system using red light in developing zebrafish embryos. Cre activity is disabled by splitting Cre and fusing the inactive halves with the Arabidopsis thaliana red light-inducible binding partners, PhyB and PIF6. In addition, PhyB-PIF6 binding requires phycocyanobilin (PCB), providing an additional layer of control. Upon exposure to red light (660 nm) illumination, the PhyB-CreC and PIF6-CreN fusion proteins come together in the presence of PCB to restore Cre activity. Red-light exposure of transgenic zebrafish embryos harboring a Cre-dependent multi-color fluorescent protein reporter (ubi:zebrabow) injected with CreLite mRNAs and PCB, resulted in Cre activity as measured by the generation of multi-spectral cell labeling in various tissues, including heart, skeletal muscle and epithelium. We show that CreLite can be used for gene manipulations in whole embryos or small groups of cells at different stages of development. CreLite provides a novel optogenetic tool for precise temporal and spatial control of gene expression in zebrafish embryos that may also be useful in cell culture, ex vivo organ culture, and other animal models for developmental biology studies.

molecular biology