Cytokinesis is essential for development and survival of all organisms. Defects in cytokinesis cause aneuploidy and genomic instability, thereby contributing to serious diseases such as cancer, neuronal disorders, and anemia. Thus, mechanistic study of cytokinesis is important not only for understanding the basic principles of a fundamental process but also for designing new strategies to treat human diseases. Cytokinesis in animal and fungal cells requires spatiotemporally coordinated functions of a contractile actomyosin ring (AMR), targeted vesicle fusion, and localized extracellular matrix (ECM) remodeling. It is much more complex than previously appreciated. In this application, we will address three major unanswered questions regarding this fundamental process using both budding yeast and mammalian cell models, with the goal of dissecting deep mechanisms in yeast and exploring evolutionary conservation in mammalian cells.
In Aim 1, we will determine the architecture of the AMR. Specifically, we will examine how myosin-II and its associated proteins such as actin and IQGAP are organized in the contractile ring from cells synchronized at cytokinesis using platinum-replica electron microscopy (PREM) coupled with immuno-gold labeling as well as super-resolution stochastic optical reconstruction microscopy (STORM).
In Aim 2, we will test our hypothesis that myosin filament assembly is regulated by heavy chain phosphorylation as well as by trans-acting factors such as IQGAP using biochemical, genetic, quantitative live imaging, and other cutting- edge imaging methods as described above.
In Aim 3, we will determine how the AMR guides exocytosis and ECM remodeling at the division site. Specifically, we will test our hypothesis that the tail of the yeast myosin-II positions and unloads vesicles from the transport machinery at the division site by interacting with the vesicle-associated guanine-nucleotide-exchange factor (GEF), Sec2, for the Rab GTPase Sec4. Then, the myosin-associated protein complex (Inn1, Hof1, and Cyk3) promotes vesicle fusion via the C2 domain of Inn1, and activates the cargo enzyme Chs2, a member of the glycosyltransferase family 2, for ECM remodeling (i.e. septum formation in yeast) via the transglutaminase-like domain of Cyk3. The discovery (Aim 1) and hypothesis-driven (Aims 2 and 3) research is expected to generate novel concepts and mechanisms of cytokinesis that are beyond specific model organisms.

Public Health Relevance

Cytokinesis is a fundamental process essential for development and survival of all organisms. Failures in cytokinesis can cause genomic instability, thereby contributing to serious diseases such as cancer, neuronal disorders, and anemia. Thus, mechanistic study of cytokinesis is important not only for understanding basic biology but also for developing new strategies to treat human diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM115420-05
Application #
9838848
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Gindhart, Joseph G
Project Start
2015-08-01
Project End
2023-07-31
Budget Start
2019-09-01
Budget End
2020-07-31
Support Year
5
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Oh, Younghoon; Schreiter, Jennifer H; Okada, Hiroki et al. (2017) Hof1 and Chs4 Interact via F-BAR Domain and Sel1-like Repeats to Control Extracellular Matrix Deposition during Cytokinesis. Curr Biol 27:2878-2886.e5
Okada, Satoshi; Lee, Mid Eum; Bi, Erfei et al. (2017) Probing Cdc42 Polarization Dynamics in Budding Yeast Using a Biosensor. Methods Enzymol 589:171-190
Okada, S; Wloka, C; Bi, E (2017) Analysis of protein dynamics during cytokinesis in budding yeast. Methods Cell Biol 137:25-45
Yang, Li; Wang, Wei-Hua; Qiu, Wei-Lin et al. (2017) A single-cell transcriptomic analysis reveals precise pathways and regulatory mechanisms underlying hepatoblast differentiation. Hepatology 66:1387-1401
Bhavsar-Jog, Yogini P; Bi, Erfei (2017) Mechanics and regulation of cytokinesis in budding yeast. Semin Cell Dev Biol 66:107-118
Ong, K; Svitkina, T; Bi, E (2016) Visualization of in vivo septin ultrastructures by platinum replica electron microscopy. Methods Cell Biol 136:73-97