FY2016 Annual Report

Developmental Neurobiology Unit
Associate Professor Ichiro Masai

Abstract

The vertebrate neural retina is derived from the ventral region of the forebrain. In this region, six classes of neurons differentiate and form the neural circuit underlying visual transduction. Thus, the retina provides an excellent model for studying cell differentiation and neural circuit formation in the vertebrate brain. Furthermore, more than one hundred hereditary retinal diseases causing photoreceptor degeneration have been identified in humans. Understanding the pathological processes of photoreceptor degeneration is an important issue from a medical perspective. We are currently investigating three research topics: (1) photoreceptor degeneration, (2) the role of microglia in retinal development and degeneration, and (3) lens development. 

1. Staff

  • Dr. Yuko Nishiwaki, Group leader
  • Dr. Sachihiro Suzuki, Staff scientist (–August 2016)
  • Dr. Maria Iribarne, Postdoctral scholar
  • Dr. Akane Hagiwara, Postdoctral scholar (–March 2017)
  • Dr. Chong Yee Hang, Postdoctral scholar (HFSP) (February 2017–) 
  • Dr. Yuki Sugiyama, Staff scientist (March 2017–)
  • Ms. Nishtha Ranawat, phD student
  • Mr. Yutaka Kojima, Technician
  • Ms. Miyuki Suenaga, Technician (–December 2016)
  • Mr. Kevin Jeff Liner, Technician
  • Ms. Lina Koronfel, Technician (–August 2016)
  • Ms. Kimberlie Ward, Technician (September 2016–)
  • Dr. Tetsuya Harakuni, Technician (October 2016–)
  • Dr. Hiroshi Izumi, Technician (December 2016–)
  • Ms. Kazumi Toguchi, Research Assistant (–March 2017)
  • Ms. Moe Inafuku, Research Assistant
  • Ms. Chitose Mizuta, Research Assistant
  • Ms. Rui Inoue, Research Assistant
  • Ms. Ayako Nakama, Research Assistant
  • Ms. Madoka Makiya, Research Assistant (tempo) (July 2016–)
  • Ms. Maki Ishikawa, Research Assistant (tempo) (January 2017–)
  • Ms. Satsuki Asato, Research Assistant (February 2017–)
  • Ms Ayako Gima, Research Administrator/Secretary

2. Collaborations

2.1 In vivo functional analysis of hypoxia response genes using the zebrafish retina

  • Description: 
  • Type of collaboration: Joint research agreement
  • Researchers: Dr. Ichiro Masai (Developmental neurobiology unit, OIST), Dr. Masayuki Matsushita (Department of Medicine, Ryukyu University)

3. Activities and Findings

3.1 Mechanism underlying photoreceptor degeneration

3.1.1 Mechanism of BNip1-mediated photoreceptor degeneration

Mitochondria-dependent apoptosis is promoted by pro-apoptotic Bcl2 family proteins, Bax, and inhibited by anti-apoptotic Bcl2 family proteins, Bcl2. BH3-only proteins promote apoptosis by modulating the balance between pro-apoptotic and anti-apoptotic Bcl2 proteins. We previously found that a BH3-only protein, BNip1, mediates thecoa-mediated photoreceptor apoptosis. BNip1 is a component of the syntaxin 18 SNARE complex, and the syntaxin 18 SNARE complex regulates retrograde transport from the Golgi apparatus to the ER. b-SNAP promotes the recycle of vesicular fusion machinery, SNARE, by disassembling the cis-SNARE complex generated by vesicular fusion. Thus, it is likely that the syntaxin 18 cis-SNARE complex is accumulated in thecoamutant. We found that failed disassembly of the syntaxin 18 cis-SNARE complex activates the BNip1-dependent photoreceptor apoptosis in the coamutant. These data suggest that the syntaxin 18 cis-SNARE complex monitors vesicular fusion competence and that BNip1 transforms vesicular fusion defects into apoptosis. From these data, we propose that BNip1 functions as an alarm that monitors abnormalities of vesicular fusion and transforms vesicular fusion defects into apoptosis in photoreceptors. 

3.1.2 Mechanism of AIPL1-mediated photoreceptor degeneration 

The photoreceptor is the neuron in charge for detecting light, and transforms this information into an electrical response. This process is known as phototransduction with the cGMP and Ca2+as the second messenger in this pathway. It has been proposed that many mutations in the components of the phototransduction produce abnormal level of cGMP and Ca2+, and finally trigger the photoreceptor death. However, the molecular mechanisms underlying the cell death remain unclear. In humans, more than a hundred genes associated with inherited photoreceptor degeneration have been mapped (RetNet,https://sph.uth.edu/Retnet).

In our lab, using different zebrafish mutant with a defect in the vision we study the degeneration process in the retina. gold rushis one of these zebrafish strain mutants, which show no photopic visual response, and slow progressive cone-specific photoreceptor degeneration. We found that the gold rushmutant gene encodes aryl hydrocarbon receptor interacting protein-like 1 (AIPL1). AIPL1 is expressed in photoreceptors and required for stability and membrane anchoring of a phototransduction molecule, cGMP-phosphodiesterase 6 (PDE6). We found that AIPL1 is an important protein not only for the phototransduction, but also for survival of cone photoreceptors in zebrafish. We previously reported that photopic vision and cone photoreceptor survival are affected in the eclipse mutant, a cone-specific PDE6 mutant (PDE6c). PDE6c has a central role in the metabolism of cGMP. Both zebrafish mutant strain, gold rush and eclipse, show a very similar cone-specific degeneration. When, we examined PDE6c functions in gold rush mutant by western blot analysis we found that the protein level of PDE6c was absent. We also found that cGMP level was higher in gold rush mutant than wild-type, and that gold rush mutation genetically enhanced photoreceptor degeneration in eclipsemutant. These data suggest that AIPL1 is required for PDE6 functions in zebrafish cone photoreceptors. 

Currently, we are trying to understand the role of the metabolism of the cGMP in the degeneration process using zebrafish phototransduction mutants. We expected that these findings would help to establish neuroprotection strategies for patients who are suffering photoreceptor degeneration.

3.2 Mechanism underlying lens development

In vertebrates, lens epithelial cells cover the anterior half of the lens fiber core. During development, lens epithelial cells proliferate, move posteriorly, and differentiate into lens fiber cells after passing through the equator. To elucidate mechanisms underlying lens epithelial cell movement, we conducted time-lapse imaging of zebrafish lens epithelium. Lens epithelial cells do not intermingle but maintain their relative positions during development. Cell division induces epithelial rearrangement, which subsequently promotes cell movement toward the equator. These data suggest that cell division is the major driving force for cell movement. In zebrafish, E-cadherin is expressed in lens epithelium, whereas N-cadherin is required for lens fiber growth. E-cadherin reduced lens epithelial cell movement, whereas N-cadherin enhanced it. Laser ablation experiments revealed that lens epithelium is governed by pulling tension, which is modulated by these cadherins. Thus, cell division and cadherin-mediated adhesion regulate lens epithelial cell movement via modulation of epithelial tension.

 

4. Publications

4.1 Journals

  1. Mochizuki, T., Luo, Y. J., Tsai, H. F., Hagiwara, A., and Masai, I. (2017) Cell division and cadherin-mediated adhesion regulates lens epithelial cell movement in zebrafish. Development144, 708-719. 

4.2 Books and other one-time publications

Nothing to report

4.3 Oral and Poster Presentations

Oral, International conference

  1. Nishiwaki, Y., Suenaga, M., Araragi, M., and Masai, I. ER-resident BH3-only protein, BNip1, induces apoptosis in response to excessive activation of vesicular transport in zebrafish photoreceptor, in XVII International Symposium on Retinal Degeneration, Kyoto, Japan. (19-24 Sept 2016).
  2. Masai, I., Spatial regulation of lens fiber differentiation, in 23rdEast Asia Joint Symposium, Taipei, Taiwan (18-20 Oct 2016). 
  3. Hagiwara, A., Mochizuki, T., Kojima, Y., Nishiwaki, Y., and Masai, I. Mechanism of ectopic lens fiber differentiation in response to early endosome trafficking defects inISER (XXII Biennial Meeting of International Society for Eye Research), Tokyo, Japan (25-29 Sept 2016)

Poster, international conferences

  1. Masai, I., Hagiwara, A, Mochizuki, T., Kojima, Y., and Nishiwaki, Y. VPS45 mutation induces ectopic lens fiber differentiation in the lens epithelium through the activation of TGF-bsignaling, inISER (XXII Biennial Meeting of International Society for Eye Research), Tokyo, Japan (25-29 Sept 2016)
  2. Nishiwaki, Y., Suenaga, S., Araragi, M., and Masai, I.  An ER-resident BH3-only protein, BNip1, induces apoptosis in response to excessive vesicular transport during photoreceptor differentiation in ISER (XXII Biennial Meeting of International Society for Eye Research), Tokyo, Japan (25-29 Sept 2016)
  3. Nishiwaki, Y., Suenaga, S., Araragi, M., and Masai, I.  Mechanism that links vesicular fusion defects and apoptosis in photoreceptor, in The12th International Conference on Zebrafish Development (The Allied Genetics2016 Conference),Orland, USA (13-17 July, 2016).
  4. Iribarne, M., Nishiwaki, Y., and Masai, I. Mutation of AIPL1 produces cone photoreceptor degeneration through a dysfunction of PDE6c and GC in zebrafish, in XVII International Symposium on Retinal Degeneration, Kyoto, Japan. (19-24 Sept 2016).

Invited talks

  1. Masai, I., Mochizuki, T., Luo, Y.-J., Tsai, H.-F., Masai, I. Cell division and cadherin-mediated adhesion regulate lens epithelial cell movement in zebrafish, in Crystalline Lens 2016 at Wakayama: Basic and Clinical Aspect, Wakayama Medical University, Japan (4-5 April, 2016). 

Oral, domestic conferences

  1. Suzuki, S., Wong, R., and Masai, I. Mechanisms underlying cone photoreceptor specification and patterning in zebrafish in JSDB Special Symposium: Frontier of Developmental Biology Hosted by JSDB,Tokyo, Japan (2 June, 2016).
  2. Nishiwaki, Y., Suenaga, S., Araragi, M., and Masai, I.  Mechanism that links vesicular fusion defects and apoptosis in photoreceptor, in The 39thAnnual meeting of the Japan Neuroscience Society, Yokohama, Japan (20-22 July, 2016). 
  3. Hagiwara, A., Mochizuki, T., Kojima, Y., Nishiwaki, Y. Yamaguchi, M. and Masai, I.  Induction mechanism of ectopic lens fiber differentiation by early endosomal trafficking defects, in The 9thRetina research meeting, Tokyo, Japan (10 Dec, 2016). 

Poster, domestic conferences

  1. Nishiwaki, Y., Suenaga, S., Araragi, M., and Masai, I.  Mechanism that links vesicular fusion defects and apoptosis in photoreceptor, in JSDB Special Symposium: Frontier of Developmental Biology Hosted by JSDB,Tokyo, Japan (2 June, 2016).
  2. Ranawat, N., Nishiwaki, Y. and Masai, I.  Brain colonization of microglia precursors during development and their role in neurodegeneration of zebrafish prim1 mutant, in JSDB Special Symposium: Frontier of Developmental Biology Hosted by JSDB,Tokyo, Japan (2 June, 2016).
  3. Kinoshita, N., Huang, A, McHugh, T., Suzuki, S., Masai, I., Miyawaki, A., and Shimogori, T.  Delineation of neuronal connectivity employing intercellular GFP reconstitution method: GRAPHIC, in The 39thAnnual meeting of the Japan Neuroscience Society, Yokohama, Japan (20-22 July, 2016). 
  4. Nishiwaki, Y., Suenaga, S., Araragi, M., and Masai, I.  Mechanism that links vesicular fusion defects and apoptosis in photoreceptor, in The 39thAnnual meeting of the Japan Society of Molecular Biology, Yokohama, Japan (30 Nov-2 Dec, 2016). 
  5. Hagiwara, A., Mochizuki, T., Kojima, Y., Nishiwaki, Y. and Masai, I.  VSP45-dependent early endosomal trafficking mechanism that regulates lens fiber differentiation in zebrafish, in The 39thAnnual meeting of the Japan Society of Molecular Biology, Yokohama, Japan (30 Nov-2 Dec, 2016). 
  6. Ranawat, N., Nishiwaki, Y. and Masai, I.  Brain colonization of microglia precursors during development and their role in neurodegeneration of zebrafish prim1 mutant, in The 39thAnnual meeting of the Japan Society of Molecular Biology, Yokohama, Japan (30 Nov-2 Dec, 2016). 

5. Intellectual Property Rights and Other Specific Achievements

Funding

KAKENHI (grants from the Ministry of Education, Science and Sport/JSPS)

Yuko Nishiwaki: Scientific research C (Kiban C) (2014–2016)

Sachihiro Suzuki: Scientific research C (Kiban C) (2015–2017)

HFSP Research grant

Ichiro Masai: Co-Investigator of HFSP Research grant (Principal Investigator: Greg Stephens)

6. Meetings and Events

6.1 Seminars

  • Speaker: Dr. Teruhiro Okuyama
  • Affiliation:  Massachusetts Institute of Technology
  • Title: “Manipulation of Social memory stored in the hippocampus”.  
  • Date: March 6, 2017
  • Venue: OIST Campus Lab1

6.2 OIST course

  • Title: Developmental Neurobiology Course 2016
  • Co-organizer:Yoko Yazaki-Sugiyama (OIST), David L. van Vactor (Harvard Medical School, OIST), Tomomi Shimogori (RIKEN BSI)
  • Dates:23 July – 8 August 2016
  • Place:OIST campus
  • Speakers: Rosalind Segal (Harvard Medical School), Yimin Zou (University of California, San Diego), Su Guo (UCSF), Takaki Miyata (Nagoya Univ.), HIsashi Umemori (Harvard Medical School), Tomomi Shimogori (RIKEN BSI), David L. van Vactor (Harvard Medical School, OIST), Hiroshi Kohsaka (University of Tokyo), Thomas Clandinin (Stanford), Yoko Yazaki-Sugiyama (OIST), Minoru Saitoe (Tokyo Metropolitan Institute of Medical Science), Naoshige Uchida (Harvard Univ.), Richard Mooney (Duke Univ.), Nirao Shah (UCSF), Atsushi Miyawaki (RIKEN BSI), Herwig Baier (Max Planck Institute of Neurobiology), Ichiro Masai (OIST), Tsuyoshi Miyakawa (Fujita Health Univ.), Yakako Morimoto (Tokyo Univ. Pharmacy and Life Sci.)

7. Other

Nothing to report.