RZZ and Mad1 dynamics in Drosophila mitosis

Chromosome Research : an International Journal on the Molecular, Supramolecular and Evolutionary Aspects of Chromosome Biology
Lénaïg DéfachellesR Karess

Abstract

The presence or absence of Mad1 at kinetochores is a major determinant of spindle assembly checkpoint (SAC) activity, the surveillance mechanism that delays anaphase onset if one or more kinetochores remain unattached to spindle fibers. Among the factors regulating the levels of Mad1 at kinetochores is the Rod, Zw10, and Zwilch (RZZ) complex, which is required for Mad1 recruitment through a mechanism that remains unknown. The relative dynamics and interactions of Mad1 and RZZ at kinetochores have not been extensively investigated, although Mad1 has been reported to be stably recruited to unattached kinetochores. In this study, we directly compare Mad1-green fluorescent protein (GFP) turnover dynamics on unattached Drosophila kinetochores with that of RZZ, tagged either with GFP-Rod or GFP-Zw10. We find that nearly 40 % of kinetochore-bound Mad1 has a significant dynamic component, turning over with a half-life of 12 s. RZZ in contrast is essentially stable on unattached kinetochores. In addition, we report that a fraction of RZZ and Mad1 can co-immunoprecipitate, indicating that the genetically determined recruitment hierarchy (in which Mad1 depends on RZZ) may reflect a physical association of the two complexes.

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Citations

Jul 1, 2020·Biology of the Cell·Alexandra Menant, Roger E Karess
Jun 6, 2018·Frontiers in Cell and Developmental Biology·Yibo LuoSong-Tao Liu
Aug 21, 2019·Frontiers in Cell and Developmental Biology·Shijie LiuBrian Storrie
Feb 8, 2019·BioEssays : News and Reviews in Molecular, Cellular and Developmental Biology·Junbin QianMathieu Bollen
Jul 11, 2019·Nucleic Acids Research·Fengjie YuanYing Zhao

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Methods Mentioned

BETA
immunoprecipitation
co-immunoprecipitation
Fluorescence
confocal microscopy
fluorescence recovery after photobleaching
transgenic
co-IP

Software Mentioned

R
Mascot
Cell
Proteome Discoverer
ImageJ
Origin
BestGene

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