PTPA variants and impaired PP2A activity in early-onset parkinsonism with intellectual disability.

The protein phosphatase 2A complex (PP2A), the major Ser/Thr phosphatase in the brain, is involved in a number of signalling pathways and functions, including the regulation of crucial proteins for neurodegeneration, such as alpha-synuclein, tau and LRRK2. Here, we report the identification of variants in the PTPA/PPP2R4 gene, encoding a major PP2A activator, in two families with early-onset parkinsonism and intellectual disability. We carried out clinical studies and genetic analyses, including genome-wide linkage analysis, whole-exome sequencing, and Sanger sequencing of candidate variants. We next performed functional studies on the disease-associated variants in cultured cells and knock-down of ptpa in Drosophila melanogaster. We first identified a homozygous PTPA variant, c.893T>G (p.Met298Arg), in patients from a South African family with early-onset parkinsonism and intellectual disability. Screening of a large series of additional families yielded a second homozygous variant, c.512C>A (p.Ala171Asp), in a Libyan family with a similar phenotype. Both variants co-segregate with disease in the respective families. The affected subjects display juvenile-onset parkinsonism and intellectual disability. The motor symptoms were responsive to treatment with levodopa and deep brain stimulation of the subthalamic nucleus. In overexpression studies, both the PTPA p.Ala171Asp and p.Met298Arg variants were associated with decreased PTPA RNA stability and decreased PTPA protein levels; the p.Ala171Asp variant additionally displayed decreased PTPA protein stability. Crucially, expression of both variants was associated with decreased PP2A complex levels and impaired PP2A phosphatase activation. PTPA orthologue knock-down in Drosophila neurons induced a significant impairment of locomotion in the climbing test. This defect was age-dependent and fully reversed by L-DOPA treatment. We conclude that bi-allelic missense PTPA variants associated with impaired activation of the PP2A phosphatase cause autosomal recessive early-onset parkinsonism with intellectual disability. Our findings might also provide new insights for understanding the role of the PP2A complex in the pathogenesis of more common forms of neurodegeneration.

Altering the Temporal Regulation of One Transcription Factor Drives Evolutionary Trade-Offs between Head Sensory Organs.

Size trade-offs of visual versus olfactory organs is a pervasive feature of animal evolution. This could result from genetic or functional constraints. We demonstrate that head sensory organ size trade-offs in Drosophila are genetically encoded and arise through differential subdivision of the head primordium into visual versus non-visual fields. We discover that changes in the temporal regulation of the highly conserved eyeless/Pax6 gene expression during development is a conserved mechanism for sensory trade-offs within and between Drosophila species. We identify a natural single nucleotide polymorphism in the cis-regulatory region of eyeless in a binding site of its repressor Cut that is sufficient to alter its temporal regulation and eye size. Because eyeless/Pax6 is a conserved regulator of head sensory placode subdivision, we propose that its temporal regulation is key to define the relative size of head sensory organs.

Architecture of the primary taste center of Drosophila melanogaster larvae.

A simple nervous system combined with stereotypic behavioral responses to tastants, together with powerful genetic and molecular tools, have turned Drosophila larvae into a very promising model for studying gustatory coding. Using the Gal4/UAS system and confocal microscopy for visualizing gustatory afferents, we provide a description of the primary taste center in the larval central nervous system. Essentially, gustatory receptor neurons target different areas of the subesophageal ganglion (SOG), depending on their segmental and sensory organ origin. We define two major and two smaller subregions in the SOG. One of the major areas is a target of pharyngeal sensilla, the other one receives inputs from both internal and external sensilla. In addition to such spatial organization of the taste center, circumstantial evidence suggests a subtle functional organization: aversive and attractive stimuli might be processed in the anterior and posterior part of the SOG, respectively. Our results also suggest less coexpression of gustatory receptors than proposed in prior studies. Finally, projections of putative second-order taste neurons seem to cover large areas of the SOG. These neurons may thus receive multiple gustatory inputs. This suggests broad sensitivity of secondary taste neurons, reminiscent of the situation in mammals.

Respective roles of the DRL receptor and its ligand WNT5 in Drosophila mushroom body development.

In recent decades, Drosophila mushroom bodies (MBs) have become a powerful model for elucidating the molecular mechanisms underlying brain development and function. We have previously characterized the derailed (drl; also known as linotte) receptor tyrosine kinase as an essential component of adult MB development. Here we show, using MARCM clones, a non-cell-autonomous requirement for the DRL receptor in MB development. This result is in accordance with the pattern of DRL expression, which occurs throughout development close to, but not inside, MB cells. While DRL expression can be detected within both interhemispheric glial and commissural neuronal cells, rescue of the drl MB defects appears to involve the latter cellular type. The WNT5 protein has been shown to act as a repulsive ligand for the DRL receptor in the embryonic central nervous system. We show here that WNT5 is required intrinsically within MB neurons for proper MB axonal growth and probably interacts with the extrinsic DRL receptor in order to stop axonal growth. We therefore propose that the neuronal requirement for both proteins defines an interacting network acting during MB development.

DmOAZ, the unique Drosophila melanogaster OAZ homologue is involved in posterior spiracle development.

In this paper, we study DmOAZ, the unique Drosophila melanogaster homologue of the OAZ zinc finger protein family. We show partial conservation of the zinc finger organization between DmOAZ and the vertebrate members of this family. We determine the exon/intron structure of the dmOAZ gene and deduce its open reading frame. Reverse transcriptase-polymerase chain reaction analysis shows that dmOAZ is transcribed throughout life. In the embryo, strongest DmOAZ expression is observed in the posterior spiracles. We suggest that dmOAZ acts as a secondary target of the Abd-B gene in posterior spiracle development, downstream of cut and ems. In a newly created loss-of-function mutant, dmOAZ ( 93 ), the "filzkörper" part of the posterior spiracles, is indeed structurally abnormal. The dmOAZ ( 93 ) mutant is a larval lethal, a phenotype that may be linked to the spiracular defect. Given the dmOAZ ( 93 ) mutant as a new tool, the fruit fly may provide an alternative model for analyzing in vivo the functions of OAZ family members.

Role of proneural genes in the formation of the larval olfactory organ of Drosophila.

In this paper, we address the role of proneural genes in the formation of the dorsal organ in the Drosophila larva. This organ is an intricate compound comprising the multineuronal dome-the exclusive larval olfactory organ-and a number of mostly gustatory sensilla. We first determine the numbers of neurons and of the different types of accessory cells in the dorsal organ. From these data, we conclude that the dorsal organ derives from 14 sensory organ precursor cells. Seven of them appear to give rise to the dome, which therefore may be composed of seven fused sensilla, whereas the other precursors produce the remaining sensilla of the dorsal organ. By a loss-of-function approach, we then analyze the role of atonal, amos, and the achaete-scute complex (AS-C), which in the adult are the exclusive proneural genes required for chemosensory organ specification. We show that atonal and amos are necessary and sufficient in a complementary way for four and three of the sensory organ precursors of the dome, respectively. AS-C, on the other hand, is implicated in specifying the non-olfactory sensilla, partially in cooperation with atonal and/or amos. Similar links for these proneural genes with olfactory and gustatory function have been established in the adult fly. However, such conserved gene function is not trivial, given that adult and larval chemosensory organs are anatomically very different and that the development of adult olfactory sensilla involves cell recruitment, which is unlikely to play a role in dome formation.

Integration of complex larval chemosensory organs into the adult nervous system of Drosophila.

The sense organs of adult Drosophila, and holometabolous insects in general, derive essentially from imaginal discs and hence are adult specific. Experimental evidence presented here, however, suggests a different developmental design for the three largely gustatory sense organs located along the pharynx. In a comprehensive cellular analysis, we show that the posteriormost of the three organs derives directly from a similar larval organ and that the two other organs arise by splitting of a second larval organ. Interestingly, these two larval organs persist despite extensive reorganization of the pharynx. Thus, most of the neurons of the three adult organs are surviving larval neurons. However, the anterior organ includes some sensilla that are generated during pupal stages. Also, we observe apoptosis in a third larval pharyngeal organ. Hence, our experimental data show for the first time the integration of complex, fully differentiated larval sense organs into the nervous system of the adult fly and demonstrate the embryonic origin of their neurons. Moreover, they identify metamorphosis of this sensory system as a complex process involving neuronal persistence, generation of additional neurons and neuronal death. Our conclusions are based on combined analysis of reporter expression from P[GAL4] driver lines, horseradish peroxidase injections into blastoderm stage embryos, cell labeling via heat-shock-induced flip-out in the embryo, bromodeoxyuridine birth dating and staining for programmed cell death. They challenge the general view that sense organs are replaced during metamorphosis.