Research Article: The Bicoid Stability Factor Controls Polyadenylation and Expression of Specific Mitochondrial mRNAs in Drosophila melanogaster

Date Published: October 13, 2011

Publisher: Public Library of Science

Author(s): Ana Bratic, Anna Wredenberg, Sebastian Grönke, James B. Stewart, Arnaud Mourier, Benedetta Ruzzenente, Christian Kukat, Rolf Wibom, Bianca Habermann, Linda Partridge, Nils-Göran Larsson, Gregory S. Barsh

Abstract: The bicoid stability factor (BSF) of Drosophila melanogaster has been reported to be present in the cytoplasm, where it stabilizes the maternally contributed bicoid mRNA and binds mRNAs expressed from early zygotic genes. BSF may also have other roles, as it is ubiquitously expressed and essential for survival of adult flies. We have performed immunofluorescence and cell fractionation analyses and show here that BSF is mainly a mitochondrial protein. We studied two independent RNAi knockdown fly lines and report that reduced BSF protein levels lead to a severe respiratory deficiency and delayed development at the late larvae stage. Ubiquitous knockdown of BSF results in a severe reduction of the polyadenylation tail lengths of specific mitochondrial mRNAs, accompanied by an enrichment of unprocessed polycistronic RNA intermediates. Furthermore, we observed a significant reduction in mRNA steady state levels, despite increased de novo transcription. Surprisingly, mitochondrial de novo translation is increased and abnormal mitochondrial translation products are present in knockdown flies, suggesting that BSF also has a role in coordinating the mitochondrial translation in addition to its role in mRNA maturation and stability. We thus report a novel function of BSF in flies and demonstrate that it has an important intra-mitochondrial role, which is essential for maintaining mtDNA gene expression and oxidative phosphorylation.

Partial Text: The maternal to zygotic transition, during which control of development shifts from maternally contributed mRNAs to genes expressed in the zygote, is of considerable interest. The maternally contributed bicoid mRNA, encoding a protein important for formation of anterior body patterning, is dependent on regulatory mechanisms controlling the cytoplasmic stability and localization of the mRNA in the zygote. The bicoid stability factor (BSF) is thought to be involved in this process as it binds the 3′UTR of the bicoid mRNA [1]. Mutation of the BSF binding site leads to reduced abundance of bicoid mRNA, whereas a P element insertion mutation that leads to a drastic reduction in BSF protein levels does not affect the abundance or distribution of endogenous bicoid mRNA [1]. In another study, BSF was reported to have a role in regulation of early zygotic genes by binding a short consensus sequence in the 5′UTR of genes expressed in the early zygote [2]. It was also reported that BSF is essential, as a P element insertion in the bsf open reading frame is lethal in the homozygous form [2]. However, homozygous bicoid mutant flies are viable and BSF therefore is likely to have an additional role besides regulating bicoid expression [2]. Characterization of global gene expression patterns in flies have shown that BSF is ubiquitously expressed in adults, further indicating that the role of BSF may not be limited to embryogenesis [3]. Studies of the subcellular localization of BSF have shown that it is present in cytoplasmic particles in oocytes and surrounding nurse cells [1] and in the cytoplasm and nucleus in early embryos [2]. Bioinformatics analyses suggest that BSF has some homology to the mammalian LRPPRC protein [1], [4], which belongs to the pentatricopeptide repeat class of proteins [5]. The LRPPRC protein has been reported to have roles in cytoplasmic RNA transport [6]–[8] and nuclear transcription [9], [10], but its main localization is in the mitochondrial matrix [4], [10], [11], where it has been suggested to stabilize mitochondrial transcripts [12].

BSF has previously been suggested to stabilize cytoplasmic mRNAs in oocytes and early zygotic cells during the first few hours of fly embryogenesis [1], [2]. However, the ubiquitous expression of the BSF RNA [3] and the punctuate cytoplasmic localization in flies has prompted us to re-investigate the function of BSF in the fly. Surprisingly, we were able to demonstrate that the BSF protein is mainly localized to mitochondria, where it controls polyadenylation of specific mitochondrial mRNAs. Further, BSF plays a key role in the regulation of mtDNA gene expression, by coordinating mitochondrial translation in flies.