Cup is essential foroskarmRNA translational repression during earlyDrosophilaoogenesis

Abstract

The proper timing of mRNA translation is crucial across many biological systems for processes such as intercellular communication, body pattern formation, and morphogenesis. The main D. melanogaster posterior determinant, oskar, is maternally transcribed, but only translated when properly localized at the oocyte's posterior cortex. Bruno 1 and Cup are two effector proteins known to participate in multiple aspects of oskar mRNA regulation. Current model describes a mechanism in which Bruno 1 is necessary for Cup's recruitment to oskar mRNA, and Bruno 1 is indispensable for its translational repression. Here, we reveal that the Bruno 1-Cup interaction, as well as their interdependent influence on each other's mRNA and protein expression, lead to precise oskar mRNA regulation during early oogenesis. We show that these factors stably associate with the oskar mRNA in vivo, but surprisingly, Bruno 1's stable association with oskar mRNA depends on Cup, while Bruno 1 is not necessary for Cup association to oskar mRNA. During early oogenesis, Cup, not Bruno 1, is the essential factor for oskar mRNA repression. Cup is a crucial P-body member that maintains proper P-body morphology during oogenesis, as well as it is necessary for oskar mRNA's association with P-bodies, thus driving the translational repression and stability of oskar mRNA. Our experimental results collectively suggest a regulatory mechanism where a feedback loop between Bruno 1 and Cup coordinates oskar mRNA regulation in the egg chamber allowing for proper development to occur.

The role of ER exit sites in maintaining P-body organization and transmitting ER stress response duringDrosophila melanogasteroogenesis

“…In sec16 RNAi egg chambers, Me31B-GFP condensates were 37% smaller and 28% more spherical compared to condensates in mCherry RNAi egg chambers (Fig. 3D and 3E), indicating that Me31B-GFP condensates are either unable to fully form, or maintain, proper condensate state (35,42). Despite many P-bodies being smaller than those in control knockdowns, there was a population of larger condensates.…”

“…As P-bodies host multiple mRNA processes, we wanted to ascertain if these processes were compromised in the absence of ERES. We focused our studies on oskar mRNA as it forms a well-studied mRNP and is known to localize into P-bodies (42,63). First, we investigated ERES's effect on translational repression by expressing endogenous Oskar-GFP in a sar1 RNAi background.…”

“…4D, S4B, and S4C). We next visualized ATP synthase mRNA, a housekeeping transcript that does not associate with P-bodies, and oskar mRNA, a known P-body member, in sar1 RNAi egg chambers (42,50). Notably, we found that the expression level and distribution of ATP synthase mRNA was unaffected, while oskar mRNA no longer formed large puncta in the nurse cell cytoplasm and was mislocalized to the anterior of the oocyte.…”