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The Batten disease Palmitoyl Protein Thioesterase 1 gene regulates neural specification and axon connectivity during Drosophila embryonic development.

Chu-LaGraff Q, Blanchette C, O'Hern P, Denefrio C - PLoS ONE (2010)

Bottom Line: Ppt1 embryos display numerous neural defects ranging from abnormal cell fate specification in a number of identified precursor lineages in the CNS, missing and disorganized neurons, faulty motoneuronal axon trajectory, and discontinuous, misaligned, and incorrect midline crossings of the longitudinal axon bundles of the ventral nerve cord.Defects in the PNS include a decreased number of sensory neurons, disorganized chordotonal neural clusters, and abnormally shaped neurons with aberrant dendritic projections.These results indicate that Ppt1 is essential for proper neuronal cell fates and organization; and to establish the local environment for proper axon guidance and fasciculation.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Union College, Schenectady, New York, United States of America. chulagrq@union.edu

ABSTRACT
Palmitoyl Protein Thioesterase 1 (PPT1) is an essential lysosomal protein in the mammalian nervous system whereby defects result in a fatal pediatric disease called Infantile Neuronal Ceroids Lipofuscinosis (INCL). Flies bearing mutations in the Drosophila ortholog Ppt1 exhibit phenotypes similar to the human disease: accumulation of autofluorescence deposits and shortened adult lifespan. Since INCL patients die as young children, early developmental neural defects due to the loss of PPT1 are postulated but have yet to be elucidated. Here we show that Drosophila Ppt1 is required during embryonic neural development. Ppt1 embryos display numerous neural defects ranging from abnormal cell fate specification in a number of identified precursor lineages in the CNS, missing and disorganized neurons, faulty motoneuronal axon trajectory, and discontinuous, misaligned, and incorrect midline crossings of the longitudinal axon bundles of the ventral nerve cord. Defects in the PNS include a decreased number of sensory neurons, disorganized chordotonal neural clusters, and abnormally shaped neurons with aberrant dendritic projections. These results indicate that Ppt1 is essential for proper neuronal cell fates and organization; and to establish the local environment for proper axon guidance and fasciculation. Ppt1 function is well conserved from humans to flies; thus the INCL pathologies may be due, in part, to the accumulation of various embryonic neural defects similar to that of Drosophila. These findings may be relevant for understanding the developmental origin of neural deficiencies in INCL.

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Ppt1 mutants exhibit abnormal RP2 axon trajectories.S16 ventral nerve cord stained with anti-β-gal to detect RN2-tau-lacZ expressed in aCC, pCC and RP2 neurons. (A) Wild type RN2-tau-lacZ pattern showing RP2 neuron and its axon trajectory (white arrow); and aCC/pCC neurons (arrowhead). (B–D). Ppt1A179T; RN2-Gal4:UAS-tauLacZ embryos display a loss of RP2 motoneurons (asterisks) and aCC/pCC neurons (arrowheads) in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. Many hemisegments have normal RP2 axon trajectory (white arrow) while others do not (black arrow). The RP2 in panel C shows an abnormal axon trajectory projecting posteriorly instead of anteriorly. Panel E is an enlarged subset of B. Anterior, up; caret, midline.
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pone-0014402-g003: Ppt1 mutants exhibit abnormal RP2 axon trajectories.S16 ventral nerve cord stained with anti-β-gal to detect RN2-tau-lacZ expressed in aCC, pCC and RP2 neurons. (A) Wild type RN2-tau-lacZ pattern showing RP2 neuron and its axon trajectory (white arrow); and aCC/pCC neurons (arrowhead). (B–D). Ppt1A179T; RN2-Gal4:UAS-tauLacZ embryos display a loss of RP2 motoneurons (asterisks) and aCC/pCC neurons (arrowheads) in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. Many hemisegments have normal RP2 axon trajectory (white arrow) while others do not (black arrow). The RP2 in panel C shows an abnormal axon trajectory projecting posteriorly instead of anteriorly. Panel E is an enlarged subset of B. Anterior, up; caret, midline.

Mentions: EVE is also essential for normal axonal guidance and innervation of the RP2 motoneuron [19] thus EVE can provide clues as to whether the observed alterations in cell fate lead to faulty axon pathfinding and innervation. To verify the mis-specification of neuronal cell fates in the EVE+ lineages as well as to assess the RP2 axonal projections, Df(1)446-20 and Ppt1A179T flies were crossed to lines bearing either the UAS-tau-LacZ or UAS-CD8-GFP reporter (RN2-Gal4:UAS-CD8-GFP or RN2-GAL4:UAS-tau-LacZ) driven by the RP2/aCC/pCC-specific Gal4 driver (RN2-Gal4) to clearly mark the cell bodies and axons of aCC and pCC, and RP2 neurons [20]. Anti- β-gal staining and GFP visualization of embryos from these crosses confirmed EVE staining results. Ppt1;RN2-Gal4:UAS-CD8-GFP and Ppt1; RN2-Gal4:UAS-tauLacZ embryos display a loss of either tauLacZ or GFP-expressing RP2s and aCC/pCC neurons in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. In many Ppt1 mutants, the remaining RP2 motoneuron exhibits abnormal axon trajectory (Figure 3). Normally at late embryogenesis, the RP2 motoneuron extends its axon anteriorly toward the ipsilateral neighboring anterior segment to join the intersegmental nerve (ISN) to exit the CNS and eventually synapsing onto the dorsal muscle (Figure 3A). In many Ppt1A179T; UAS-tauLacZ embryos, the RP2 neurons extend its axons in the wrong direction-posteriorly to join the posterior ipsilateral ISN instead (Figure 3D) while other RP2 axons project contralaterally toward the midline, stalled, and never exit the CNS (Figure 3E). These embryos also display a loss of either aCC or pCC neurons in many hemisegments (Figure 3B–E). Collectively, these studies indicated that the loss of Ppt1 caused abnormal neuronal cell fate specification and defective axon pathfinding.


The Batten disease Palmitoyl Protein Thioesterase 1 gene regulates neural specification and axon connectivity during Drosophila embryonic development.

Chu-LaGraff Q, Blanchette C, O'Hern P, Denefrio C - PLoS ONE (2010)

Ppt1 mutants exhibit abnormal RP2 axon trajectories.S16 ventral nerve cord stained with anti-β-gal to detect RN2-tau-lacZ expressed in aCC, pCC and RP2 neurons. (A) Wild type RN2-tau-lacZ pattern showing RP2 neuron and its axon trajectory (white arrow); and aCC/pCC neurons (arrowhead). (B–D). Ppt1A179T; RN2-Gal4:UAS-tauLacZ embryos display a loss of RP2 motoneurons (asterisks) and aCC/pCC neurons (arrowheads) in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. Many hemisegments have normal RP2 axon trajectory (white arrow) while others do not (black arrow). The RP2 in panel C shows an abnormal axon trajectory projecting posteriorly instead of anteriorly. Panel E is an enlarged subset of B. Anterior, up; caret, midline.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3008717&req=5

pone-0014402-g003: Ppt1 mutants exhibit abnormal RP2 axon trajectories.S16 ventral nerve cord stained with anti-β-gal to detect RN2-tau-lacZ expressed in aCC, pCC and RP2 neurons. (A) Wild type RN2-tau-lacZ pattern showing RP2 neuron and its axon trajectory (white arrow); and aCC/pCC neurons (arrowhead). (B–D). Ppt1A179T; RN2-Gal4:UAS-tauLacZ embryos display a loss of RP2 motoneurons (asterisks) and aCC/pCC neurons (arrowheads) in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. Many hemisegments have normal RP2 axon trajectory (white arrow) while others do not (black arrow). The RP2 in panel C shows an abnormal axon trajectory projecting posteriorly instead of anteriorly. Panel E is an enlarged subset of B. Anterior, up; caret, midline.
Mentions: EVE is also essential for normal axonal guidance and innervation of the RP2 motoneuron [19] thus EVE can provide clues as to whether the observed alterations in cell fate lead to faulty axon pathfinding and innervation. To verify the mis-specification of neuronal cell fates in the EVE+ lineages as well as to assess the RP2 axonal projections, Df(1)446-20 and Ppt1A179T flies were crossed to lines bearing either the UAS-tau-LacZ or UAS-CD8-GFP reporter (RN2-Gal4:UAS-CD8-GFP or RN2-GAL4:UAS-tau-LacZ) driven by the RP2/aCC/pCC-specific Gal4 driver (RN2-Gal4) to clearly mark the cell bodies and axons of aCC and pCC, and RP2 neurons [20]. Anti- β-gal staining and GFP visualization of embryos from these crosses confirmed EVE staining results. Ppt1;RN2-Gal4:UAS-CD8-GFP and Ppt1; RN2-Gal4:UAS-tauLacZ embryos display a loss of either tauLacZ or GFP-expressing RP2s and aCC/pCC neurons in many hemisegments; and disorganized cellular arrangements of the remaining LacZ-positive neurons. In many Ppt1 mutants, the remaining RP2 motoneuron exhibits abnormal axon trajectory (Figure 3). Normally at late embryogenesis, the RP2 motoneuron extends its axon anteriorly toward the ipsilateral neighboring anterior segment to join the intersegmental nerve (ISN) to exit the CNS and eventually synapsing onto the dorsal muscle (Figure 3A). In many Ppt1A179T; UAS-tauLacZ embryos, the RP2 neurons extend its axons in the wrong direction-posteriorly to join the posterior ipsilateral ISN instead (Figure 3D) while other RP2 axons project contralaterally toward the midline, stalled, and never exit the CNS (Figure 3E). These embryos also display a loss of either aCC or pCC neurons in many hemisegments (Figure 3B–E). Collectively, these studies indicated that the loss of Ppt1 caused abnormal neuronal cell fate specification and defective axon pathfinding.

Bottom Line: Ppt1 embryos display numerous neural defects ranging from abnormal cell fate specification in a number of identified precursor lineages in the CNS, missing and disorganized neurons, faulty motoneuronal axon trajectory, and discontinuous, misaligned, and incorrect midline crossings of the longitudinal axon bundles of the ventral nerve cord.Defects in the PNS include a decreased number of sensory neurons, disorganized chordotonal neural clusters, and abnormally shaped neurons with aberrant dendritic projections.These results indicate that Ppt1 is essential for proper neuronal cell fates and organization; and to establish the local environment for proper axon guidance and fasciculation.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Union College, Schenectady, New York, United States of America. chulagrq@union.edu

ABSTRACT
Palmitoyl Protein Thioesterase 1 (PPT1) is an essential lysosomal protein in the mammalian nervous system whereby defects result in a fatal pediatric disease called Infantile Neuronal Ceroids Lipofuscinosis (INCL). Flies bearing mutations in the Drosophila ortholog Ppt1 exhibit phenotypes similar to the human disease: accumulation of autofluorescence deposits and shortened adult lifespan. Since INCL patients die as young children, early developmental neural defects due to the loss of PPT1 are postulated but have yet to be elucidated. Here we show that Drosophila Ppt1 is required during embryonic neural development. Ppt1 embryos display numerous neural defects ranging from abnormal cell fate specification in a number of identified precursor lineages in the CNS, missing and disorganized neurons, faulty motoneuronal axon trajectory, and discontinuous, misaligned, and incorrect midline crossings of the longitudinal axon bundles of the ventral nerve cord. Defects in the PNS include a decreased number of sensory neurons, disorganized chordotonal neural clusters, and abnormally shaped neurons with aberrant dendritic projections. These results indicate that Ppt1 is essential for proper neuronal cell fates and organization; and to establish the local environment for proper axon guidance and fasciculation. Ppt1 function is well conserved from humans to flies; thus the INCL pathologies may be due, in part, to the accumulation of various embryonic neural defects similar to that of Drosophila. These findings may be relevant for understanding the developmental origin of neural deficiencies in INCL.

Show MeSH
Related in: MedlinePlus