Drosophila histone locus bodies form by hierarchical recruitment of components (original) (raw)
Drosophila Histone Locus Body assembly and function involves multiple interactions
2020
The histone locus body (HLB) assembles at replication-dependent (RD) histone loci and concentrates factors required for RD histone mRNA biosynthesis. The D. melanogaster genome has a single locus comprised of ∼100 copies of a tandemly arrayed repeat unit containing one copy of each of the 5 RD histone genes. To determine sequence elements required for D. melanogaster HLB formation and histone gene expression, we used transgenic gene arrays containing 12 copies of the histone repeat unit that functionally complement loss of the ∼200 endogenous RD histone genes. A 12x histone gene array in which all H3-H4 promoters were replaced with H2a-H2b promoters does not form an HLB or express high levels of RD histone mRNA in the presence of the endogenous histone genes. In contrast, this same transgenic array is active in HLB assembly and RD histone gene expression in the absence of the endogenous RD histone genes and rescues the lethality caused by homozygous deletion of the RD histone locus....
Developmental and Cell Cycle Regulation of the Drosophila Histone Locus Body
Molecular Biology of the Cell, 2007
Cyclin E/Cdk2 is necessary for replication-dependent histone mRNA biosynthesis, but how it controls this process in early development is unknown. We show that in Drosophila embryos the MPM-2 monoclonal antibody, raised against a phosphoepitope from human mitotic cells, detects Cyclin E/Cdk2-dependent nuclear foci that colocalize with nascent histone transcripts. These foci are coincident with the histone locus body (HLB), a Cajal body-like nuclear structure associated with the histone locus and enriched in histone pre-mRNA processing factors such as Lsm11, a core component of the U7 small nuclear ribonucleoprotein. Using MPM-2 and anti-Lsm11 antibodies, we demonstrate that the HLB is absent in the early embryo and occurs when zygotic histone transcription begins during nuclear cycle 11. Whereas the HLB is found in all cells after its formation, MPM-2 labels the HLB only in cells with active Cyclin E/Cdk2. MPM-2 and Lsm11 foci are present in embryos lacking the histone locus, and MPM-2 foci are present in U7 mutants, which cannot correctly process histone pre-mRNA. These data indicate that MPM-2 recognizes a Cdk2-regulated protein that assembles into the HLB independently of histone mRNA biosynthesis. HLB foci are present in histone deletion embryos, although the MPM-2 foci are smaller, and some Lsm11 foci are not associated with MPM-2 foci, suggesting that the histone locus is important for HLB integrity.
Genes & Development, 2009
We generated mutant alleles of Drosophila melanogaster in which expression of the linker histone H1 can be down-regulated over a wide range by RNAi. When the H1 protein level is reduced to ;20% of the level in wildtype larvae, lethality occurs in the late larval -pupal stages of development. Here we show that H1 has an important function in gene regulation within or near heterochromatin. It is a strong dominant suppressor of position effect variegation (PEV). Similar to other suppressors of PEV, H1 is simultaneously involved in both the repression of euchromatic genes brought to the vicinity of pericentric heterochromatin and the activation of heterochromatic genes that depend on their pericentric localization for maximal transcriptional activity. Studies of H1-depleted salivary gland polytene chromosomes show that H1 participates in several fundamental aspects of chromosome structure and function. First, H1 is required for heterochromatin structural integrity and the deposition or maintenance of major pericentric heterochromatin-associated histone marks, including H3K9Me 2 and H4K20Me 2 . Second, H1 also plays an unexpected role in the alignment of endoreplicated sister chromatids. Finally, H1 is essential for organization of pericentric regions of all polytene chromosomes into a single chromocenter. Thus, linker histone H1 is essential in Drosophila and plays a fundamental role in the architecture and activity of chromosomes in vivo.
Molecular biology of the cell, 2015
Nuclear bodies (NBs) are structures that concentrate proteins, RNAs, and ribonucleoproteins that perform functions essential to gene expression. How NBs assemble is not well understood. We studied the Drosophila histone locus body (HLB), a NB that concentrates factors required for histone mRNA biosynthesis at the replication-dependent histone gene locus. We coupled biochemical analysis with confocal imaging of both fixed and live tissues to demonstrate that the Drosophila Multi-Sex Combs (Mxc) protein contains multiple domains necessary for HLB assembly. An important feature of this assembly process is the self-interaction of Mxc via two conserved N-terminal domains: a LisH domain and a novel SIF (Self Interaction Facilitator) domain immediately downstream of the LisH domain. Molecular modeling suggests that the LisH and SIF domains directly interact, and mutation of either the LisH or SIF domains severely impairs Mxc function in vivo resulting in reduced histone mRNA accumulation. ...
Genes & Development, 2009
We generated mutant alleles of Drosophila melanogaster in which expression of the linker histone H1 can be down-regulated over a wide range by RNAi. When the H1 protein level is reduced to ;20% of the level in wildtype larvae, lethality occurs in the late larval -pupal stages of development. Here we show that H1 has an important function in gene regulation within or near heterochromatin. It is a strong dominant suppressor of position effect variegation (PEV). Similar to other suppressors of PEV, H1 is simultaneously involved in both the repression of euchromatic genes brought to the vicinity of pericentric heterochromatin and the activation of heterochromatic genes that depend on their pericentric localization for maximal transcriptional activity. Studies of H1-depleted salivary gland polytene chromosomes show that H1 participates in several fundamental aspects of chromosome structure and function. First, H1 is required for heterochromatin structural integrity and the deposition or maintenance of major pericentric heterochromatin-associated histone marks, including H3K9Me 2 and H4K20Me 2 . Second, H1 also plays an unexpected role in the alignment of endoreplicated sister chromatids. Finally, H1 is essential for organization of pericentric regions of all polytene chromosomes into a single chromocenter. Thus, linker histone H1 is essential in Drosophila and plays a fundamental role in the architecture and activity of chromosomes in vivo.
Genetics, 2006
Chromatin packaging directly influences gene programming as it permits only certain portions of the genome to be activated in any given developmental stage, cell, and tissue type. Histone acetyltransferases (HATs) are a key class of chromatin regulatory proteins that mediate such developmental chromatin control; however, their specific roles during multicellular development remain unclear. Here, we report the first isolation and developmental characterization of a Drosophila HAT gene (Dmel \TIP60) that is the homolog of the human HAT gene TIP60. We show that Dmel \TIP60 is differentially expressed during Drosophila development, with transcript levels significantly peaking during embryogenesis. We further demonstrate that reducing endogenous Dmel \TIP60 expression in Drosophila embryonic cells by RNAi results in cellular defects and lethality. Finally, using a GAL4-targeted RNAi system in Drosophila, we show that ubiquitous or mesoderm/muscle-specific reduction of Dmel \TIP60 expression results in lethality during fly development. Our results suggest a mechanism for HAT regulation involving developmental control of HAT expression profiles and show that Dmel \TIP60 is essential for multicellular development. Significantly, our inducible and targeted HAT knockdown system in Drosophila now provides a powerful tool for effectively studying the roles of TIP60 in specific tissues and cell types during development.
PLoS ONE, 2009
Background: Metazoan replication-dependent histone mRNAs terminate in a conserved stem-loop structure rather than a polyA tail. Formation of this unique mRNA 39 end requires Stem-loop Binding Protein (SLBP), which directly binds histone pre-mRNA and stimulates 39 end processing. The 39 end stem-loop is necessary for all aspects of histone mRNA metabolism, including replication coupling, but its importance to organism fitness and genome maintenance in vivo have not been characterized. Methodology/Principal Findings: In Drosophila, disruption of the Slbp gene prevents normal histone pre-mRNA processing and causes histone pre-mRNAs to utilize the canonical 39 end processing pathway, resulting in polyadenylated histone mRNAs that are no longer properly regulated. Here we show that Slbp mutants display genomic instability, including loss of heterozygosity (LOH), increased presence of chromosome breaks, tetraploidy, and changes in position effect variegation (PEV). During imaginal disc growth, Slbp mutant cells show defects in S phase and proliferate more slowly than control cells. Conclusions/Significance: These data are consistent with a model in which changing the 39 end of histone mRNA disrupts normal replication-coupled histone mRNA biosynthesis and alters chromatin assembly, resulting in genomic instability, inhibition of cell proliferation, and impaired development.
Drosophila melanogaster HI histone is phosphorylated stably
Molecular and Cellular Biology
Phosphorylation of histone Hi is developmentally regulated in Drosophila spp. It cannot be detected in preblastoderm embryos or polytene salivary gland cells, but in cellular blastoderm, postblastoderm embryo, and amitotic adplt head nuclei, it occurs with a frequency of roughly 4 x i05 molecules per nucleus. We used pulse-labeling to study the relationship-between HI'$ynthesis and modification in cultured cells. These results reveal that the HI-associated phosphate is stable and siggest that Drosophila Hi is synthesized, translocated to the nuc!eus,. associated with chromatin, and then phosphorylated. Partial tryptic digestion of Drosophila HI revealed that the phosphorylation site is located within the globular, central domain of the protein. Thus, the developmentally regulated phosphorylation of Drosophila Hi presents two contrasts with previously studied HI phosphorylation. 'It is not correlated with DNA replication, and it is located in the central domain of the protein.
The histone acetyltransferase Dmel\TIP60 Is essential for multicellular development in Drosophila
2007
Combinatorial histone modifications control chromatin packaging which in turn, contributes to the precise patterning of gene expression during development. Histone acetyltransferases (HATs) are a key class of chromatin regulatory proteins that mediate such developmental chromatin control; however their specific roles during multicellular development remain unclear. Here, we report the first isolation and developmental characterization of a Drosophila HAT gene (Dmel\\\\\\\\TIP60) that is the homolog of the human HAT gene TIP60. We show that Dmel\\\\\\\\TIP60 is differentially expressed during Drosophila development, with transcript levels significantly peaking during embryogenesis. We further demonstrate that reducing endogenous Dmel\\\\\\\\TIP60 expression in Drosophila embryonic cells by RNAi results in cellular defects and lethality. Finally, we use our Drosophila GAL4 inducible Dmel\\\\\\\\TIP60 knockdown/overexpression system to explore the role of Dmel\\\\\\\\TIP60 in a wide va...
Histone locus regulation by the Drosophila dosage compensation adaptor protein CLAMP
Genes & Development
The conserved histone locus body (HLB) assembles prior to zygotic gene activation early during development and concentrates factors into a nuclear domain of coordinated histone gene regulation. Although HLBs form specifically at replication-dependent histone loci, the cis and trans factors that target HLB components to histone genes remained unknown. Here we report that conserved GA repeat cis elements within the bidirectional histone3–histone4 promoter direct HLB formation in Drosophila. In addition, the CLAMP (chromatin-linked adaptor for male-specific lethal [MSL] proteins) zinc finger protein binds these GA repeat motifs, increases chromatin accessibility, enhances histone gene transcription, and promotes HLB formation. We demonstrated previously that CLAMP also promotes the formation of another domain of coordinated gene regulation: the dosage-compensated male X chromosome. Therefore, CLAMP binding to GA repeat motifs promotes the formation of two distinct domains of coordinate...