Biology subjects

Bratu, D. P.

Publications and source records attributed to Bratu, D. P..

6 recordsLinked to original sources

bioRxiv2026

ATF4-dependent upregulation of Bruno 1 remodels P-bodies to selectively protect mRNAs during ER stress throughout Drosophila melanogaster oogenesis

P-bodies are cytoplasmic membraneless organelles involved in mRNA storage, yet their role in cellular stress responses remains poorly understood. Here, we demonstrate that P-bodies are rapidly and selectively remodeled during the early response to endoplasmic reticulum (ER) stress in D. melanogaster oogenesis, positioning them as key early stress responders. Notably, this remodeling occurs within minutes of stress induction and precedes stress granule formation. This early remodeling is characterized by changes in P-body morphology and internal organization and promotes selective mRNA regulation. Specifically, ER stress leads to the recruitment and stabilization of maternal mRNAs and those encoding P-body components, while transcripts not associated with P-bodies are degraded. These observations indicate that P-body remodeling is not merely structural but functionally linked to the selective preservation of mRNA populations during stress. Mechanistically, we find that this process is driven by transcriptional upregulation of the RNA-binding protein, Bruno 1, downstream of ATF4-dependent stress signaling, thereby establishing a direct connection between the unfolded protein response and condensate regulation. Consistent with this model, loss of Bruno 1 abolishes, whereas its overexpression enhances P-body remodeling, demonstrating that stress-induced changes in RNA binding protein levels can actively reprogram condensate properties. Together, our findings reveal that P-bodies function as dynamic, stress-responsive hubs that integrate transcriptional signaling with post-transcriptional control, enabling the selective preservation of essential mRNAs during ER stress. More broadly, this work uncovers a previously unrecognized mechanism by which stress signaling pathways reorganize cytoplasmic architecture to shape mRNA fate.

cell biology↗

bioRxiv2024

Post-transcriptional regulation of cyclin A and cyclin B mRNAs is mediated by Bruno 1 and Cup, and further fine-tuned within P-bodies

Cell cycle progression is tightly controlled by the regulated synthesis and degradation of Cyclins, such as Cyclin A and Cyclin B, which activate CDK1 to trigger mitosis. Mutations affecting Cyclin regulation are often linked to tumorigenesis, making the study of cyclin mRNA regulation critical for identifying new cancer therapies. In this study, we demonstrate via super-resolution microscopy that cyclin A and cyclin B mRNAs associate with Bruno 1 and Cup in nurse cells. The depletion of either protein leads to abnormal Cyclin A and Cyclin B protein expression and a reduction in mRNA levels for both Cyclins. We further reveal that both cyclin A and cyclin B mRNAs accumulate in P-bodies marked by Me31B. Interestingly, Me31B is not involved in regulating cyclin A mRNA, as no changes in cyclin A mRNA levels or repression are observed upon Me31B depletion. However, cyclin B mRNA shows stage-specific derepression and reduced levels when Me31B is absent. Notably, the association between cyclin B and Cup is strengthened in the absence of Me31B, indicating that this interaction occurs independently of P-bodies. These results highlight the nuanced, mRNA-specific roles of P-body condensates in post-transcriptional regulation, challenging the idea of a uniform, binary mechanism of mRNA repression in P-bodies.

cell biology↗

bioRxiv2024

The role of ER exit sites in maintaining P-body organization and transmitting ER stress response during Drosophila melanogaster oogenesis

Processing bodies (P-bodies) are cytoplasmic membrane-less organelles which host multiple mRNA processing events. While the fundamental principles of P-body organization are beginning to be elucidated in vitro, a nuanced understanding of how their assembly is regulated in vivo remains elusive. Here, we investigate the potential link between ER exit sites and P-bodies in Drosophila melanogaster egg chambers. Employing a combination of live and super-resolution imaging, we found that P-bodies associated with ER exit sites are larger and less mobile than cytoplasmic P-bodies, indicating that they constitute a distinct class of P-bodies which are more mature than their cytoplasmic counterparts. Moreover, we demonstrate that altering the composition of ER exit sites has differential effects on core P-body proteins (Me31B, Cup, and Trailer Hitch) suggesting a potential role for ER exit sites in P-body organization. We further show that in the absence of ER exit sites, P-body integrity is compromised and the stability and translational repression efficiency of the maternal mRNA, oskar, are reduced. Finally, we show that ER stress is communicated to P-bodies via ER exit sites, highlighting the pivotal role of ER exit sites as a bridge between membrane-bound and membrane-less organelles in ER stress response. Together, our data unveils the significance of ER exit sites not only in governing P-body organization, but also in facilitating inter-organellar communication during stress, potentially bearing implications for a variety of disease pathologies.

cell biology↗

bioRxiv2023

Cup is essential for oskar mRNA translational repression during early Drosophila oogenesis

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 oocytes 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 Cups 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 others 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 1s 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 mRNAs 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.

developmental biology↗

bioRxiv2018

PinMol: Python application for designing molecular beacons for live cell imaging of endogenous mRNAs

Molecular beacons are nucleic acid oligomers labeled with a fluorophore and a quencher that fold in a hairpin-shaped structure, which fluoresce only when bound to their target RNA. They are used for the visualization of endogenous mRNAs in live cells. Here, we report a Python program (PinMol) that designs molecular beacons best suited for live cell imaging by using structural information from secondary structures of the target RNA, predicted via energy minimization approaches. PinMol takes into account the accessibility of the targeted regions, as well as the inter- and intramolecular interactions of each selected probe. To demonstrate its applicability, we synthesized an oskar mRNA-specific molecular beacon (osk1236), which is selected by PinMol to target a more accessible region than a manually designed oskar-specific molecular beacon (osk2216). We previously demonstrated osk2216 to be efficient in detecting oskar mRNA in in vivo experiments. Here, we show that osk1236 outperformed osk2216 in live cell imaging experiments.

molecular biology↗

bioRxiv2018

P-bodies and the miRNA pathway regulate translational repression of bicoid mRNA during Drosophila melanogaster oogenesis

Embryonic axis patterning in Drosophila melanogaster is partly achieved by mRNAs that are maternally localized to the oocyte; the spatio-temporal regulation of these transcripts stability and translation is a characteristic feature of oogenesis. While protein regulatory factors are necessary for the translational regulation of some maternal transcripts (e.g. oskar and gurken), small RNA pathways are also known to regulate mRNA stability and translation in eukaryotes. MicroRNAs (miRNAs) are small RNA regulators of gene expression, widely conserved throughout eukaryotic genomes and essential for animal development. The main D. melanogaster anterior determinant, bicoid, is maternally transcribed, but it is not translated until early embryogenesis. We investigated the possibility that its translational repression during oogenesis is mediated by miRNA activity. We found that the bicoid 3UTR contains a highly conserved, predicted binding site for miR-305. Our studies reveal that miR-305 regulates the translation of a reporter gene containing the bicoid 3UTR in cell culture, and that miR-305 only partially contributes to bicoid mRNA translational repression during oogenesis. We also found that Processing bodies (P-bodies) in the egg chamber may play a role in stabilizing bicoid and other maternal transcripts. Here, we offer insights into the possible role of P-bodies and the miRNA pathway in the translational repression of bicoid mRNA during oogenesis.

cell biology↗