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  • 1.
    O'Farrell, Fergal
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institutet.
    A screen for mutations affecting PNS development in Drosophila identifies the trim gene, dappled2008Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    The peripheral nervous system of Drosophila melanogaster contains a variety of sense organs, ranging from the relatively simple four celled bristle organ to the more complex compound eye. The development of each organ type is well described, providing a useful backdrop for functional studies of genes acting in one or more of the many processes involved in organogenesis. We have used the bristle organ to screen for genes affecting PNS development. Two of the candidates recovered via this approach, string (stg, Drosophila cdc25, the universal regulator of the G2 to M phase mitotic transition), and dappled (dpld, a poorly described gene implicated in tumor suppression) were selected for further study. Examination of stg mis-expression phenotypes in the adult bristle organ revealed cell fate transformations corresponding to the generation of two pIIa structural precursor cells at the expense of a neural precursor cell. This transformation most reasonably resulted from an abnormally short G2 arrest, indicating that the time spent in the G2 phase is crucial to correct cell fate determination. dpld is a member of the Tripartite Motif (TRIM) superfamily, members of which are involved in diverse biological processes e.g. proliferation, apoptosis and immune response. dpld belongs to a subgroup of NHL domain containing TRIM proteins, that are known to be involved in tumor suppression. Phylogenetic analysis placed dpld in the lin-41 sub-clade of the TRIM superfamily. A combination of insilico, genetic and cell culture assay approaches showed dpld to be susceptible to miRNA regulation. As homologous genes are also miRNA regulated this regulatory mechanism may be conserved throughout this sub-clade, between vertebrates and invertebrates. Pre-existing loss of function dpld alleles were characterized, however, subsequent complementation studies revealed that characteristic aspects of the described dpld phenotype, in fact mapped outside the dpld locus, and were caused by mutations of nearby genes. The tumor-causing locus was mapped to the Cytb5 gene (mutated in both pre-existing dpld alleles), while the embryonic lethality and PNS phenotype was mapped to the scraps locus. scraps encodes for Drosophila Anillin, known to be required during cytokinesis. We provide the first characterization of scraps null alleles and detail a biased requirement for scraps within neural precursor cells of the embryonic PNS. A novel loss of function dpld allele was recovered. This mutation is lethal, however it does not have an associated tumor phenotype. This finding, together with our complementation study indicates that the existing classification of dpld as a tumor suppressor is inaccurate. Subsequent studies detail dpld requirements in the developing fly retina. There, dpld mutation resulted in excessive proliferation, while conversely, mis-expression caused a reduction. Additionally, and perhaps consequently, cell differentiation was affected. Thus, regulation of proliferation by NHL-TRIM genes seems a conserved feature. We additionally identified a novel Drosophila TRIM gene of the same class as dpld, which we have dubbed another bbox affiliate (abba), bringing the number of NHL containing TRIM genes in Drosophila to four.

  • 2.
    O'Farrell, Fergal
    et al.
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institute.
    Esfahani, Shiva Seyedoleslami
    Stockholms universitet.
    Engström, Ylva
    Stockholms universitet.
    Kylsten, Per
    Södertörns högskola, Institutionen för livsvetenskaper.
    Regulation of the Drosophila lin-41 homologue dappled by let-7 reveals conservation of a regulatory mechanism within the LIN-41 subclade2008Inngår i: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 237, nr 1, s. 196-208Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Drosophila Dappled (DPLD) is a member of the RBCC/TRIM superfamily, a protein family involved in numerous diverse processes such as developmental timing and asymmetric cell divisions. DPLD belongs to the LIN-41 subclade, several members of which are micro RNA (miRNA) regulated. We re-examined the LIN-41 subclade members and their relation to other RBCC/TRIMs and dpld paralogs, and identified a new Drosophila muscle specific RBCC/TRIM: Another B-Box Affiliate, ABBA. In silico predictions of candidate miRNA regulators of dpld identified let-7 as the strongest candidate. Overexpression of dpld led to abnormal eye development, indicating that strict regulation of dpld mRNA levels is crucial for normal eye development. This phenotype was sensitive to let-7 dosage, suggesting let-7 regulation of dpld in the eye disc. A cell-based assay verified let-7 miRNA down-regulation of dpld expression by means of its 3'-untranslated region. Thus, dpld seems also to be miRNA regulated, suggesting that miRNAs represent an ancient mechanism of LIN-41 regulation.

  • 3.
    O'Farrell, Fergal
    et al.
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institutet.
    Kylsten, Per
    Södertörns högskola, Institutionen för livsvetenskaper.
    A mis-expression study of factors affecting Drosophila PNS cell identity2008Inngår i: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 370, nr 4, s. 657-662Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Drosophila PNS sense organs arise from single sensory organ precursor (SOP) cells through a series of asymmetric divisions. In a mis-expression screen for factors affecting PNS development, we identified string and dappled as being important for the proper formation of adult external sensory (ES) organs. string is a G2 regulator. dappled has no described function but is implicated in tumorigenesis. The misexpression effect from string was analysed using timed over expression during adult ES-organ and, for comparison, embryonic Chorclotonal (Ch) organ formation. Surprisingly, string mis-expression prior to SOP division gave the greatest effect in both systems. In adult ES-organs, this lead to cell fate transformations producing structural cells, whilst in the embryo organs were lost, hence differences within the lineages exist. Mis-expression of dappled, lead to loss and duplications of entire organs in both systems, potentially affecting SOP specification, in addition to affecting neuronal guidance.

  • 4.
    O'Farrell, Fergal
    et al.
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institutet.
    Kylsten, Per
    Södertörns högskola, Institutionen för livsvetenskaper.
    Drosophila Anillin is unequally required during asymmetric cell divisions of the PNS2008Inngår i: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 369, nr 2, s. 407-413Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    During Drosophila embryogenesis, timely and orderly asymmetric cell divisions ensure the correct number of each cell type that make up the sensory organs of the larval PNS. We report a role of scraps, Drosophila Anillin, during these divisions. Anillin, a constitutive member of the contractile ring is essential for cytokinesis in Drosophila and vertebrates. During embryogenesis we find that zygotically transcribed scraps is required specifically for the unequal cell divisions, those in which cytokinesis occurs in an "off-centred" manner, of the pIIb and pIIIb neuronal precursor cells, but not the equal cell divisions of the lineage related precursor cells. Complementation and genetic rescue studies demonstrate this effect results from zygotic scraps and leads to polyploidy, ectopic mitosis, and loss of the neuronal precursor daughter cells. The net result of which is the formation of incomplete sense organs and embryonic lethality.

  • 5.
    O'Farrell, Fergal
    et al.
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institutet.
    Munoz-Alarcón, Andrés
    Södertörns högskola, Institutionen för livsvetenskaper. Karolinska Institutet.
    Georgiev, Alexander
    Stockholms universitet.
    Kylsten, Per
    Södertörns högskola, Institutionen för livsvetenskaper.
    Functional Analysis of Drosophila lin-41, dappled during adult eye developmentManuskript (preprint) (Annet vitenskapelig)
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