Pre-Fibrillar α-Synuclein Mutants Cause Parkinson's Disease-Like Non-Motor Symptoms in Drosophila

Parkinson's disease (PD) is linked to the formation of insoluble fibrillar aggregates of the presynaptic protein α-Synuclein (αS) in neurons. The appearance of such aggregates coincides with severe motor deficits in human patients. These deficits are often preceded by non-motor symptoms such as sleep-related problems in the patients. PD-like motor deficits can be recapitulated in model organisms such as Drosophila melanogaster when αS is pan-neurally expressed. Interestingly, both these deficits are more severe when αS mutants with reduced aggregation properties are expressed in flies. This indicates that that αS aggregation is not the primary cause of the PD-like motor symptoms. Here we describe a model for PD in Drosophila which utilizes the targeted expression of αS mutants in a subset of dopadecarboxylase expressing serotonergic and dopaminergic (DA) neurons. Our results show that targeted expression of pre-fibrillar αS mutants not only recapitulates PD-like motor symptoms but also the preceding non-motor symptoms such as an abnormal sleep-like behavior, altered locomotor activity and abnormal circadian periodicity. Further, the results suggest that the observed non-motor symptoms in flies are caused by an early impairment of neuronal functions rather than by the loss of neurons due to cell death

Bällchen is required for self-renewal of germline stem cells in Drosophila melanogaster

Self-renewing stem cells are pools of undifferentiated cells, which are maintained in cellular niche environments by distinct tissue-specific signalling pathways. In Drosophila melanogaster, female germline stem cells (GSCs) are maintained in a somatic niche of the gonads by BMP signalling. Here we report a novel function of the Drosophila kinase Bällchen (BALL), showing that its cell autonomous role is to maintain the self-renewing capacity of female GSCs independent of BMP signalling. ball mutant GSCs are eliminated from the niche and subsequently differentiate into mature eggs, indicating that BALL is largely dispensable for differentiation. Similar to female GSCs, BALL is required to maintain self-renewal of male GSCs, suggesting a tissue independent requirement of BALL for self-renewal of germline stem cells

Bällchen participates in proliferation control and prevents the differentiation of Drosophila melanogaster neuronal stem cells

Stem cells continuously generate differentiating daughter cells and are essential for tissue homeostasis and development. Their capacity to self-renew as undifferentiated and actively dividing cells is controlled by either external signals from a cellular environment, the stem cell niche, or asymmetric distribution of cell fate determinants during cell division. Here we report that the protein kinase Bällchen (BALL) is required to prevent differentiation as well as to maintain normal proliferation of neuronal stem cells of Drosophila melanogaster, called neuroblasts. Our results show that the brains of ball mutant larvae are severely reduced in size, which is caused by a reduced proliferation rate of the neuroblasts. Moreover, ball mutant neuroblasts gradually lose the expression of the neuroblast determinants Miranda and aPKC, suggesting their premature differentiation. Our results indicate that BALL represents a novel cell intrinsic factor with a dual function regulating the proliferative capacity and the differentiation status of neuronal stem cells during development

Drosophila Separase is required for sister chromatid separation and binds to PIM and THR

Drosophila PIM and THR are required for sister chromatid separation in mitosis and associate in vivo. Neither of these two proteins shares significant sequence similarity with known proteins. However, PIM has functional similarities with securin proteins. Like securin, PIM is degraded at the metaphase-to-anaphase transition and this degradation is required for sister chromatid separation. Securin binds and inhibits separase, a conserved cysteine endoprotease. Proteolysis of securin at the metaphase-to-anaphase transition activates separase, which degrades a conserved cohesin subunit, thereby allowing sister chromatid separation. To address whether PIM regulates separase activity or functions with THR in a distinct pathway, we have characterized a Drosophila separase homolog (SSE). SSE is an unusual member of the separase family. SSE is only about one-third the size of other separases and has a diverged endoprotease domain. However, our genetic analyses show that SSE is essential and required for sister chromatid separation during mitosis. Moreover, we show that SSE associates with both PIM and THR. Although our work shows that separase is required for sister chromatid separation in higher eukaryotes, in addition, it also indicates that the regulatory proteins have diverged to a surprising degree, particularly in Drosophila

Diverse stimuli engage different neutrophil extracellular trap pathways

Neutrophils release neutrophil extracellular traps (NETs) which ensnare pathogens and have pathogenic functions in diverse diseases. We examined the NETosis pathways induced by five stimuli; PMA, the calcium ionophore A23187, nigericin, Candida albicans and Group B Streptococcus. We studied NET production in neutrophils from healthy donors with inhibitors of molecules crucial to PMA induced NETs including protein kinase C, calcium, reactive oxygen species, the enzymes myeloperoxidase (MPO) and neutrophil elastase. Additionally, neutrophils from chronic granulomatous disease patients, carrying mutations in the NADPH oxidase complex or a MPO-deficient patient were examined. We show that PMA, C. albicans and GBS use a related pathway for NET induction whereas ionophores require an alternative pathway but that NETs produced by all stimuli are proteolytically active, kill bacteria and composed mainly of chromosomal DNA. Thus, we demonstrate that NETosis occurs through several signalling mechanisms, suggesting that extrusion of NETs is important in host defence

Age-dependent circadian defects in response to wild type αS, TP-αS, EKO/Kir2.1 and NaChBac expression in DA neurons.

<p>(<b>A</b>, <b>B</b>) Double-plotted actograms of young flies (3 days after hatching) expressing wild type αS (WT-αS) (A) and TP-αS (B) under the control of the DA neuron specific <i>TH-Gal4</i> driver. T refers to circadian periodicity which is 23.8 hrs in both cases. (<b>C</b>, <b>D</b>) Double-plotted actograms of old flies (30 days after hatching) expressing WT-αS and TP-αS. Note the different circadian periodicity in response to WT-αS (T = 23.7 hrs) and TP-αS expression (T = 26.7 hrs). (<b>E</b>, <b>F</b>) Double-plotted actogram of old flies expressing EKO/Kir2.1 (E) or NaChBac (F) under the control of the DA neuron specific <i>TH-Gal4</i> driver. Note the similar extension of the circadian periodicities (T = 27.6 hrs and T = 27.0 hrs, respectively) as observed after TP-αS expression. All experiments (n = 32–58 flies) were carried out under constant dark conditions after the animals were kept in a dark-light cycle of 12∶12 hrs. T was calculated by the Chi-squared periodogram analysis (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024701#s4" target="_blank">Materials and Methods</a>).</p