FY2013 Annual Report

Formation and Regulation of Neuronal Connectivity Unit

Visiting Professor David Van Vactor

Abstract

Regulation of gene expression is of great importance during the formation and maintenance of synaptic connections in the nervous system.  Through a number of collaborations, our unit is exploring mechanisms of transcriptome regulation in motor neuron differentiation and neuromuscular junction development.  We utilize the genetic model organism Drosophila in order to combine powerful genetic manipulation with in vivo analysis of neuronal cell biology and gene expression.  Our current projects focus on two areas: (a) the function of Survival of Motor Neurons (SMN), causal factor in the motor neuron disease Spinal Muscular Atrophy (SMA), and (b) the function of the conserved microRNA-8 in controlling early stages of synaptogenesis in Drosophila.

1. Staff

  • Dr. David Van Vactor, Professor
  • Dr. Takakazu Yokokura, Group Leader
  • Cecilia Lu, Research Scientist (STG)
  • Seiko Yoshikawa  (November, 2013 -)
  • Khulekani Mncube, Research Intern (Oct. 31, 2013 – Feb. 27, 2014)
  • Shino Fibbs, Research Administrator

2. Collaborations

  • Theme: Computational analysis of RNA splicing patterns in high-content RNAseq datasets, and conservation of RNA splicing and stability effects from Drosophila to human models of Spinal Muscular Atrophy (SMA).
    • Type of collaboration: Joint research
    • Researchers: Dr. Margarida Gama-Carvalho (University of Lisbon, Lisbon, Portugal)
  • Theme: Defining patterns of RNA splicing and stability characteristic of human IPS-derived mixed motor neuron model of Spinal Muscular Atrophy (SMA).
    • Type of collaboration: Joint research
    • Researchers: Dr. Lee Rubin (Dept. of Stem Cell and Regenerative Biology, Harvard University, Boston, MA, USA)

3. Activities and Findings

3.1 Spinal Muscular Atrophy (SMA)

Over the past year, our unit has continued to optimize deep sequencing (RNAseq) methods in collaboration with the OIST for Drosophila and Human RNA samples.  After completing a preliminary round of RNAseq for Drosophila CNS tissues wild type and mutant for SMN, we and our collaborators have analyzed these data.  The initial experiments reveal that analytical strategies will be required for quality control of RNAseq datasets, and establishing this bioinformatics process will be the subject of a joint publication with the Gama-Carvalho laboratory.  Once these methods are developed, our major focus will be to generate a second round of higher quality RNAseq for both Drosophila and Human models of SMA, and then perform a transcriptome-level analysis of steady state mRNA levels and RNA splicing patterns.  These future data will be compared to our recently published genetic interactome for SMA in Drosophila (Sen et al., 2013), as a means to determine if functional SMN interaction overlaps significantly with RNA splicing events that are SMN-dependent.  Although SMN is known to participate in a protein complex required for snRNP assembly, the source of dysfunction for motor neurons and neuromuscular synapses is unclear.

3.2 Spinal Muscular Atrophy (SMA)

Although our recent work showed that the conserved Drosophila microRNA miR-8 regulates the early refinement of motor neuron targeting through maintaining tissue-specific patterns of expression for two cell adhesion molecules, Neuroglian (Nrg) and Fasciclin-III (FasIII), this regulatory relationship is not direct, as neither Nrg or FasIII mRNAs carry miR-8 microRNA recognition elements (MREs).  In our search for intermediaries, we have identified a candidate transcription factor that is likely to be both downstream of miR-8 and upstream of Nrg.  Our analysis of this factor is consistent with a model where miR-8 tunes levels of multiple presynaptic and postsynaptic effector proteins through indirect regulation of transcription.

4. Publications

4.1 Journals

  1. J. Amaral, et al. (2014). "Quality assessment and control of issue specific RNA-seq libraries of Drosophila transgenic RNAi models." Frontiers in Genetics 5(43).
  2. Sen, A., et al. (2013). "Genetic circuitry of Survival motor neuron, the gene underlying spinal muscular atrophy." Proc Natl Acad Sci U S A 110(26): E2371-2380.

4.2 Books and Other One-Time Publications

Nothing to report.

4.3 Oral and Poster Presentations

  1. Lu, C. S. (2014). The Conserved MicroRNA miR-8 Regulates Synapse Morphogenesis. 55th Annual Drosophila Research Conference. San Diego, CA, USA.
  2. Van Vactor, D. (2013). History of Developmental Biology. Developmental Neurobiology Course 2013, Okinawa, Japan.
  3. Van Vactor, D. (2013). The Regulation of Synaptogenesis in the Drosophila Neuromuscular System. Developmental Neurobiology Course 2013, Okinawa, Japan. 
  4. Yokokura, T. (2013). Analysis of underlining molecular mechanism of SMA. 25th Takato Symposium. Ohtsu, Shiga, Japan.
  5. Yokokura, T. (2013). Finding right career path. Osaka, Tokyo, Japan.
  6. Yokokura, T. (2013). Identify genetic circuitry in which Smn is integrated. OIST, Okinawa, Japan.
  7. Yokokura, T. (2014). Defining Spinal Muscular Atrophy Gene Networds in Drosophila. Maggot Meeting Behavioral Neurogenetics of Larval Drosophila: Molecule, Circuits, Computation & Robotics, Atami, Shizuoka, Japan.
  8. Yokokura, T. (2014). Defining Spinal Muscular Atrophy Gene Networks in Drosopila. 55th Annual Drosophila Research Conference. San Diego, CA, USA.

5. Intellectual Property Rights and Other Specific Achievements

Nothing to report

6. Meetings and Events

6.1 Developmental Neurobiology Course 2013

  • Date: July 16 – August 1, 2013
  • Venue: Seaside House/OIST Campus
  • Organizers: Drs. David Van Vactor, Gordon Arbuthnott, Ichiro Masai, and Yoko Yazaki-Sugiyama
  • Speakers:
    • Prof. Takao Hensch (Dept. of Molecular and Cellular Biology, Harvard University)
    • Dr. Fumio Matsuzaki (RIKEN Center for Developmental Biology)
    • Prof. Masayuki Miura (Dept. of Genetics, Graduate School of Pharmaceutical Science, The University of Tokyo)
    • Assoc. Prof. Naoyuki Inagaki (Graduate School of Biological Sciences, Nara Institute of Science and Technology)
    • Prof. Anne Hart (Dept. of Neuroscience, Brown Research )
    • Prof. Jeff Lichtman (Dept. of Molecular and Cellular Biology, Harvard University)
    • Prof. Anthony Koleske (Biological & Biomedical Sciences, Yale University)
    • Prof. Rachel Wong (Dept. of Biological Structure, University of Washington)
    • Prof. Dan Sanes (Faculty of Arts & Science, New York University)
    • Prof. Hitoshi Sakano (Dept. of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo)
    • Prof. Akinao Nose (Dept. of Complexity Science and Engineering Formation and Function of the Neural Network, The University of Tokyo)
    • Assoc. Prof. Nirao Shah (Dept. of Anatomy, University of California San Francisco)
    • Dr. Hitoshi Okamoto (RIKEN BSI)
    • Dr. Atsushi Miyawaki (RIKEN BSI)
    • Prof. Zhigang He (Children’s Hospital Boston, Harvard Medical School)
    • Prof. Robin Ali (UCL Institute of Ophthalmology, Division of Molecular Therapy)
    • Prof. Yuichi Iino (Dept. of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo)
    • Assist. Prof. Bernd Kuhn (Optical Neuroimaging Unit, OIST)