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NuMA mediated control of cleavage furrow formation in Human mitosis

Work done in the lab of Prof. Sachin Kotak at the Department of Microbiology and Cell Biology (MCB) in the Indian Institute of Science (IISc), Bengaluru

About author

Ashwathi Rajeevan

Ashwathi Rajeevan obtained her BS-MS degree in Life Science from the Indian Institute of Science Education and Research (IISER), Bhopal. She is currently pursuing her Ph.D. under the mentorship of Prof. Sachin Kotak at the Department of Microbiology and Cell Biology at the Indian Institute of Science (IISc), Bengaluru, where she is studying the mechanisms that regulate various processes at mitotic exit like cleavage furrow formation and chromatin decondensation.

Interview

How would you explain your research outcomes to the non-scientific community?

The cell division (or mitosis) ensures accurate segregation of the genetic material and other cellular constituents from a mother cell to two daughter cells. Two vital events that occur in mitosis are the separation of the sister chromatids and the formation of the cleavage furrow that ultimately bisects the mother cell into two. The former event is regulated by the NuMA/dynein complex (NuMA-based complex), which is present in the polar membrane regions but absent at the equatorial membrane. We asked why the NuMA-based complex is excluded from the equatorial membrane and the relevance of this exclusion.

Through RNAi-mediated depletion experiments, we showed that the proteins critical for cleavage furrow formation, which include Cyk4, Mklp1, and Ect2, are critical for excluding the NuMA-based complex from the equatorial membrane. Further, we showed that the complex formed by Ect2, Cyk4, and Mklp1 (Ect2-based complex) and NuMA-based complex show mutually exclusive membrane localization (Figure 1). The Ect2-based complex at the equatorial membrane restricts the NuMA-based complex to the polar membrane, which is critical for chromosome separation in anaphase. Similarly, the polar-membrane localized NuMA-based complexes prevent the spreading of Ect2-based complexes to these regions, which ensures proper cleavage furrow formation and cell division. Overall, our work proposes that the physical crowding created by NuMA-based complexes and Ect2-based complexes on different cell membrane regions coordinates chromosome separation and cleavage furrow formation (Figure 1B).

Figure 1:NuMA/dynein and Ect2/Cyk4/Mklp1 show polarized membrane distribution in anaphase (A) Immunofluorescence analysis of HeLa cell in anaphase, stained with anti-NuMA (red) and anti-RhoA (green) antibodies. RhoA is the downstream protein of Ect2, Cyk4, and Mklp1. DNA is shown in blue. The scale bar represents 10 µm. (B) Schematic representation of the mutually exclusive membrane domains formed by two protein complexes- NuMA/dynein/dynactin at the polar membrane and Ect2/Cyk4/Mklp1 at the equatorial membrane- in mitotic cells at the anaphase stage. A quadrant part of the anaphase cell (shown on the left side) is highlighted on the right. The pink, black and green structures indicate chromosomes, cleavage furrow, and microtubules, respectively.
Figure 1:NuMA/dynein and Ect2/Cyk4/Mklp1 show polarized membrane distribution in anaphase (A) Immunofluorescence analysis of HeLa cell in anaphase, stained with anti-NuMA (red) and anti-RhoA (green) antibodies. RhoA is the downstream protein of Ect2, Cyk4, and Mklp1. DNA is shown in blue. The scale bar represents 10 µm. (B) Schematic representation of the mutually exclusive membrane domains formed by two protein complexes- NuMA/dynein/dynactin at the polar membrane and Ect2/Cyk4/Mklp1 at the equatorial membrane- in mitotic cells at the anaphase stage. A quadrant part of the anaphase cell (shown on the left side) is highlighted on the right. The pink, black and green structures indicate chromosomes, cleavage furrow, and microtubules, respectively.

How do these findings contribute to your research area?

Previous studies from our lab and other groups have shown multiple roles of NuMA at different stages of the cell cycle that collectively ensure proper cell division. However, NuMA has never been linked with cytokinesis. Through our study, we have shown a novel role of NuMA in regulating cleavage furrow formation due to its compartmentalized membrane distribution along with dynein/dynactin. Our study also shed light on how the polarized membrane distribution of multiple protein complexes can work together to regulate processes such as chromosome separation and cleavage furrow formation during cell division.

What was the exciting moment during your research?

In some way, there is always excitement in knowing the results of each experiment that we do during research. A few of these moments include obtaining colonies after molecular cloning and seeing the beautiful immunofluorescence images of the cells. Regarding this project, I was delighted to see a significantly diminished RhoA at the equatorial membrane upon NuMA depletion, suggesting our hypothesis was true.

“Our study also shed light on how the polarized membrane distribution of multiple protein complexes can work together to regulate processes such as chromosome separation and cleavage furrow formation during cell division.”

What do you hope to do next?

With the experience I gained from my doctoral research, I would definitely like to pursue a career in research. With respect to this project, we would like to dwell on the mechanisms of how these protein complexes are maintained at the cell membrane.

Where do you seek scientific inspiration from?

It is inspiring to see how various processes within a cell are so precisely coordinated and regulated to make it fully functional. My primary inspiration to pursue biology as a subject comes from all my teachers throughout my school and undergraduate studies. My mentor, Dr. Sachin Kotak, and my lab mates at IISc have inspired me throughout my Ph.D. Additionally, I am deeply motivated by the many scientific talks I have had the opportunity to attend.

How do you intend to help Indian science improve?

India has a lot of potential for quality scientific research. I believe a collective effort from the government and us researchers is required to improve Indian science. There must be more funding and international and industrial collaborations for researchers in India.

Reference

Shrividya Sana*, Ashwathi Rajeevan*, and Sachin Kotak (2022); Membrane compartmentalization of Ect2/Cyk4/Mklp1 and NuMA/dynein regulates cleavage furrow formation; J. Cell Biol. (October 2022) 221 (12). https://doi.org/10.1083/jcb.202203127

*Both authors contributed equally to this work

Copy Editor: Sukanya Madhwal

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