The Multiple Mitotic Roles of the ASPM Orthologous Proteins: Insight into the Etiology of ASPM-Dependent Microcephaly.

The () gene was discovered about 40 years ago and shown to be required for both mitotic and meiotic cell division. Subsequent studies showed that is highly conserved and that mutations in its human ortholog (; or ) are the most common cause of autosomal recessive primary microcephaly. This finding greatly stimulated research on and its fly and mouse () orthologs.

TGS1 impacts snRNA 3'-end processing, ameliorates survival motor neuron-dependent neurological phenotypes in vivo and prevents neurodegeneration.

Trimethylguanosine synthase 1 (TGS1) is a highly conserved enzyme that converts the 5'-monomethylguanosine cap of small nuclear RNAs (snRNAs) to a trimethylguanosine cap. Here, we show that loss of TGS1 in Caenorhabditis elegans, Drosophila melanogaster and Danio rerio results in neurological phenotypes similar to those caused by survival motor neuron (SMN) deficiency. Importantly, expression of human TGS1 ameliorates the SMN-dependent neurological phenotypes in both flies and worms, revealing that TGS1 can partly counteract the effects of SMN deficiency.

Genetic Control of Kinetochore-Driven Microtubule Growth in Mitosis.

Centrosome-containing cells assemble their spindles exploiting three main classes of microtubules (MTs): MTs nucleated by the centrosomes, MTs generated near the chromosomes/kinetochores, and MTs nucleated within the spindle by the augmin-dependent pathway. Mammalian and cells lacking the centrosomes generate MTs at kinetochores and eventually form functional bipolar spindles. However, the mechanisms underlying kinetochore-driven MT formation are poorly understood.

TEM Imaging of Membrane Choreography During Mitosis of Drosophila Tissue Culture Cells.

Schneider 2 (S2) cells are one of the most widely used Drosophila cell lines, and are specifically suitable for genetic dissection of biological processes by RNA interference. We have recently developed a method that allows an easy preparation of samples for transmission electron microscopy (TEM) analysis of S2 cells. This method is based on the collection and pelleting of the cells in test tubes, followed by fixation and staining of pellets in the same tubes.

The S-adenosylmethionine analog sinefungin inhibits the trimethylguanosine synthase TGS1 to promote telomerase activity and telomere lengthening.

Mutations in many genes that control the expression, the function, or the stability of telomerase cause telomere biology disorders (TBDs), such as dyskeratosis congenita, pulmonary fibrosis, and aplastic anemia. Mutations in a subset of the genes associated with TBDs cause reductions of the telomerase RNA moiety hTR, thus limiting telomerase activity. We have recently found that loss of the trimethylguanosine synthase TGS1 increases both hTR abundance and telomerase activity and leads to telomere elongation.

Moonlighting in Mitosis: Analysis of the Mitotic Functions of Transcription and Splicing Factors.

Moonlighting proteins can perform one or more additional functions besides their primary role. It has been posited that a protein can acquire a moonlighting function through a gradual evolutionary process, which is favored when the primary and secondary functions are exerted in different cellular compartments. Transcription factors (TFs) and splicing factors (SFs) control processes that occur in interphase nuclei and are strongly reduced during cell division, and are therefore in a favorable situation to evolve moonlighting mitotic functions.

Absence of SCAPER causes male infertility in humans and by modulating microtubule dynamics during meiosis.

Background: Mutation () have been found across ethnicities and have been shown to cause variable penetrance of an array of pathological traits, including intellectual disability, retinitis pigmentosa and ciliopathies.

Methods: Human clinical phenotyping, surgical testicular sperm extraction and testicular tissue staining. Generation and analysis of () ( orthologue) CAS9-knockout lines.

Intimate functional interactions between TGS1 and the Smn complex revealed by an analysis of the Drosophila eye development.

Trimethylguanosine synthase 1 (TGS1) is a conserved enzyme that mediates formation of the trimethylguanosine cap on several RNAs, including snRNAs and telomerase RNA. Previous studies have shown that TGS1 binds the Survival Motor Neuron (SMN) protein, whose deficiency causes spinal muscular atrophy (SMA). Here, we analyzed the roles of the Drosophila orthologs of the human TGS1 and SMN genes.

Loss of Human TGS1 Hypermethylase Promotes Increased Telomerase RNA and Telomere Elongation.

Biogenesis of the human telomerase RNA (hTR) involves a complex series of posttranscriptional modifications, including hypermethylation of the 5' mono-methylguanosine cap to a tri-methylguanosine cap (TMG). How the TMG cap affects hTR maturation is unknown. Here, we show that depletion of trimethylguanosine synthase 1 (TGS1), the enzyme responsible for cap hypermethylation, increases levels of hTR and telomerase.

Morgana is an Hsp90-interacting protein with a direct role in microtubule polymerisation.

Morgana (Mora, also known as CHORD in flies) and its mammalian homologue, called CHORDC1 or CHP1, is a highly conserved cysteine and histidine-rich domain (CHORD)-containing protein that has been proposed to function as an Hsp90 co-chaperone. Morgana deregulation promotes carcinogenesis in both mice and humans while, in , loss of causes lethality and a complex mitotic phenotype that is rescued by a human transgene. Here, we show that Mora localises to mitotic spindles and co-purifies with the Hsp90-R2TP-TTT supercomplex and with additional well-known Hsp90 co-chaperones.

RNAi-mediated depletion of the NSL complex subunits leads to abnormal chromosome segregation and defective centrosome duplication in Drosophila mitosis.

The Drosophila Nonspecific Lethal (NSL) complex is a major transcriptional regulator of housekeeping genes. It contains at least seven subunits that are conserved in the human KANSL complex: Nsl1/Wah (KANSL1), Dgt1/Nsl2 (KANSL2), Rcd1/Nsl3 (KANSL3), Rcd5 (MCRS1), MBD-R2 (PHF20), Wds (WDR5) and Mof (MOF/KAT8). Previous studies have shown that Dgt1, Rcd1 and Rcd5 are implicated in centrosome maintenance.

Publisher Correction: The role of Patronin in Drosophila mitosis.

During production of the original article [1], there was a technical error that resulted in author corrections not being rendered in the PDF version of the article.

The role of Patronin in Drosophila mitosis.

Background: The calmodulin-regulated spectrin-associated proteins (CAMSAPs) belong to a conserved protein family, which includes members that bind the polymerizing mcrotubule (MT) minus ends and remain associated with the MT lattice formed by minus end polymerization. Only one of the three mammalian CAMSAPs, CAMSAP1, localizes to the mitotic spindle but its function is unclear. In Drosophila, there is only one CAMSAP, named Patronin.

Correction to: DNA copy number evolution in Drosophila cell lines.

Following publication of the original article [1], the authors reported the following errors.

Splicing factors Sf3A2 and Prp31 have direct roles in mitotic chromosome segregation.

Several studies have shown that RNAi-mediated depletion of splicing factors (SFs) results in mitotic abnormalities. However, it is currently unclear whether these abnormalities reflect defective splicing of specific pre-mRNAs or a direct role of the SFs in mitosis. Here, we show that two highly conserved SFs, Sf3A2 and Prp31, are required for chromosome segregation in both and human cells.

Phenotypic characterization of diamond (dind), a Drosophila gene required for multiple aspects of cell division.

Many genes are required for the assembly of the mitotic apparatus and for proper chromosome behavior during mitosis and meiosis. A fruitful approach to elucidate the mechanisms underlying cell division is the accurate phenotypic characterization of mutations in these genes. Here, we report the identification and characterization of diamond (dind), an essential Drosophila gene required both for mitosis of larval brain cells and for male meiosis.

Ultrastructural analysis of mitotic Drosophila S2 cells identifies distinctive microtubule and intracellular membrane behaviors.

Background: S2 cells are one of the most widely used Drosophila melanogaster cell lines. A series of studies has shown that they are particularly suitable for RNAi-based screens aimed at the dissection of cellular pathways, including those controlling cell shape and motility, cell metabolism, and host-pathogen interactions. In addition, RNAi in S2 cells has been successfully used to identify many new mitotic genes that are conserved in the higher eukaryotes, and for the analysis of several aspects of the mitotic process.

Mice with reduced expression of the telomere-associated protein Ft1 develop p53-sensitive progeroid traits.

Human AKTIP and mouse Ft1 are orthologous ubiquitin E2 variant proteins involved in telomere maintenance and DNA replication. AKTIP also interacts with A- and B-type lamins. These features suggest that Ft1 may be implicated in aging regulatory pathways.

The Hybrid Incompatibility Genes and Are Required for Sister Chromatid Detachment During Anaphase but Not for Centromere Function.

Crosses between females and males produce hybrid sons that die at the larval stage. This hybrid lethality is suppressed by loss-of-function mutations in the () or in the () genes. Previous studies have shown that Hmr and Lhr interact with heterochromatin proteins and suppress expression of transposable elements within It also has been proposed that Hmr and Lhr function at the centromere.

The Drosophila orthologue of the INT6 onco-protein regulates mitotic microtubule growth and kinetochore structure.

INT6/eIF3e is a highly conserved component of the translation initiation complex that interacts with both the 26S proteasome and the COP9 signalosome, two complexes implicated in ubiquitin-mediated protein degradation. The INT6 gene was originally identified as the insertion site of the mouse mammary tumor virus (MMTV), and later shown to be involved in human tumorigenesis. Here we show that depletion of the Drosophila orthologue of INT6 (Int6) results in short mitotic spindles and deformed centromeres and kinetochores with low intra-kinetochore distance.

Drosophila Male Meiosis.

In Drosophila males, there is no synaptonemal complex and recombination does not occur. Thus, Drosophila male meiosis is a good model system for the analysis of achiasmate chromosome segregation. In addition, due to their large size, the meiotic spindles of Drosophila males are an excellent system for mutational dissection of the mechanisms of spindle assembly.

Citron Kinase Deficiency Leads to Chromosomal Instability and TP53-Sensitive Microcephaly.

Mutations in citron (CIT), leading to loss or inactivation of the citron kinase protein (CITK), cause primary microcephaly in humans and rodents, associated with cytokinesis failure and apoptosis in neural progenitors. We show that CITK loss induces DNA damage accumulation and chromosomal instability in both mammals and Drosophila. CITK-deficient cells display "spontaneous" DNA damage, increased sensitivity to ionizing radiation, and defective recovery from radiation-induced DNA lesions.

A Role for the Twins Protein Phosphatase (PP2A-B55) in the Maintenance of Genome Integrity.

The protein phosphatase 2A (PP2A) is a conserved heterotrimeric enzyme that regulates several cellular processes including the DNA damage response and mitosis. Consistent with these functions, PP2A is mutated in many types of cancer and acts as a tumor suppressor. In mammalian cells, PP2A inhibition results in DNA double strand breaks (DSBs) and chromosome aberrations (CABs).

The Drosophila telomere-capping protein Verrocchio binds single-stranded DNA and protects telomeres from DNA damage response.

Drosophila telomeres are sequence-independent structures maintained by transposition to chromosome ends of three specialized retroelements rather than by telomerase activity. Fly telomeres are protected by the terminin complex that includes the HOAP, HipHop, Moi and Ver proteins. These are fast evolving, non-conserved proteins that localize and function exclusively at telomeres, protecting them from fusion events.