Publications by authors named "Jamuna S Sreeja"

6 Publications

  • Page 1 of 1

α-Fodrin in Cytoskeletal Organization and the Activity of Certain Key Microtubule Kinesins.

Genes (Basel) 2021 May 17;12(5). Epub 2021 May 17.

Cancer Research Program, Rajiv Gandhi Centre for Biotechnology, University of Kerala, Thiruvananthapuram 695014, India.

Cortical cytoskeletal proteins are significant in controlling various cellular mechanisms such as migration, cell adhesion, intercellular attachment, cellular signaling, exo- and endocytosis and plasma membrane integrity, stability and flexibility. Our earlier studies involving in vitro and ex vivo approaches led us to identify certain undiscovered characteristics of α-fodrin, a prominent cortical protein. The conventional functions attributed to this protein mainly support the plasma membrane. In the present study, we utilized a global protein expression analysis approach to detect underexplored functions of this protein. We report that downregulation of α-fodrin in glioblastoma cells, U-251 MG, results in upregulation of genes affecting the regulation of the cytoskeleton, cell cycle and apoptosis. Interestingly, certain key microtubule kinesins such as KIF23, KIF2B and KIF3C are downregulated upon α-fodrin depletion, as validated by real-time PCR studies.
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http://dx.doi.org/10.3390/genes12050750DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8156673PMC
May 2021

A Fresh Look at the Structure, Regulation, and Functions of Fodrin.

Mol Cell Biol 2020 08 14;40(17). Epub 2020 Aug 14.

Cancer Research Program-III, Rajiv Gandhi Centre for Biotechnology, University of Kerala, Thiruvananthapuram, India

Fodrin and its erythroid cell-specific isoform spectrin are actin-associated fibrous proteins that play crucial roles in the maintenance of structural integrity in mammalian cells, which is necessary for proper cell function. Normal cell morphology is altered in diseases such as various cancers and certain neuronal disorders. Fodrin and spectrin are two-chain (αβ) molecules that are encoded by paralogous genes and share many features but also demonstrate certain differences. Fodrin (in humans, typically a heterodimer of the products of the SPTAN1 and SPTBN1 genes) is expressed in nearly all cell types and is especially abundant in neuronal tissues, whereas spectrin (in humans, a heterodimer of the products of the SPTA1 and SPTB1 genes) is expressed almost exclusively in erythrocytes. To fulfill a role in such a variety of different cell types, it was anticipated that fodrin would need to be a more versatile scaffold than spectrin. Indeed, as summarized here, domains unique to fodrin and its regulation by Ca, calmodulin, and a variety of posttranslational modifications (PTMs) endow fodrin with additional specific functions. However, how fodrin structural variations and misregulated PTMs may contribute to the etiology of various cancers and neurodegenerative diseases needs to be further investigated.
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http://dx.doi.org/10.1128/MCB.00133-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7431051PMC
August 2020

α-Fodrin is required for the organization of functional microtubules during mitosis.

Cell Cycle 2019 Oct 27;18(20):2713-2726. Epub 2019 Aug 27.

Cancer Research Program-III, Rajiv Gandhi Centre for Biotechnology, University of Kerala , Thiruvananthapuram , India.

The cytoskeleton protein α-fodrin plays a major role in maintaining structural stability of membranes. It was also identified as part of the brain γ-tubulin ring complex, the major microtubule nucleator. Here, we investigated the requirement of α-fodrin for microtubule spindle assembly during mitotic progression. We found that α-fodrin depletion results in abnormal mitosis with uncongressed chromosomes, leading to prolonged activation of the spindle assembly checkpoint and a severe mitotic delay. Further, α-fodrin repression led to the formation of shortened spindles with unstable kinetochore-microtubule attachments. We also found that the mitotic kinesin CENP-E had reduced levels at kinetochores to likely account for the chromosome misalignment defects in α-fodrin-depleted cells. Importantly, we showed these cells to exhibit reduced levels of detyrosinated α-tubulin, which primarily drives CENP-E localization. Since proper microtubule dynamics and chromosome alignment are required for completion of normal mitosis, this study reveals an unforeseen role of α-fodrin in regulating mitotic progression. Future studies on these lines of observations should reveal important mechanistic insight for fodrin's involvement in cancer.
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http://dx.doi.org/10.1080/15384101.2019.1656476DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6773225PMC
October 2019

Binding of alpha-fodrin to gamma-tubulin accounts for its role in the inhibition of microtubule nucleation.

FEBS Lett 2019 06 17;593(11):1154-1165. Epub 2019 May 17.

Cancer Research Programme, Rajiv Gandhi Centre for Biotechnology, Trivandrum, India.

Non-erythroid spectrin or fodrin is present as part of the γ-tubulin ring complex (γ-TuRC) in brain tissue and brain derived cells. Here, we show that fodrin, which is otherwise known for providing structural support to the cell membrane, interacts directly with γ-tubulin within the γ-TuRC through a GRIP2-like motif. Turbidometric analysis of microtubule polymerization with nucleation-potent γ-TuRC isolated from HEK-293 cells that lack fodrin and the γ-TuRC from goat brain that contains fodrin shows inefficiency of the latter to promote nucleation. The involvement of fodrin was confirmed by the reduction in the microtubule polymerization efficiency of HEK-293 derived γ-TuRCs upon addition of purified brain fodrin. Thus, the interaction of fodrin with gamma-tubulin is responsible for its inhibitory effect on γ-tubulin mediated microtubule nucleation.
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http://dx.doi.org/10.1002/1873-3468.13425DOI Listing
June 2019

+TIP EB1 downregulates paclitaxel‑induced proliferation inhibition and apoptosis in breast cancer cells through inhibition of paclitaxel binding on microtubules.

Int J Oncol 2015 Jan 7;46(1):133-46. Epub 2014 Oct 7.

School of Biology, Indian Institute of Science Education and Research, CET Campus, Thiruvananthapuram 695016, Kerala, India.

Microtubule plus‑end‑binding protein (+TIP) EB1 has been shown to be upregulated in breast cancer cells and promote breast tumor growth in vivo. However, its effect on the cellular actions of microtubule‑targeted drugs in breast cancer cells has remained poorly understood. By using cellular and biochemical assays, we demonstrate that EB1 plays a critical role in regulating the sensitivity of breast cancer cells to anti‑microtubule drug, paclitaxel (PTX). Cell viability assays revealed that EB1 expression in the breast cancer cell lines correlated with the reduction of their sensitivity to PTX. Knockdown of EB1 by enzymatically‑prepared siRNA pools (esiRNAs) increased PTX‑induced cytotoxicity and sensitized cells to PTX‑induced apoptosis in three breast cancer cell lines, MCF‑7, MDA MB‑231 and T47D. Apoptosis was associated with activation of caspase‑9 and an increase in the cleavage of poly(ADP‑ribose) polymerase (PARP). p53 and Bax were upregulated and Bcl2 was downregulated in the EB1‑depleted PTX‑treated MCF‑7 cells, indicating that the apoptosis occurs via a p53‑dependent pathway. Following its upregulation, the nuclear accumulation of p53 and its association with cellular microtubules were increased. EB1 depletion increased PTX‑induced microtubule bundling in the interphase cells and induced formation of multiple spindle foci with abnormally elongated spindles in the mitotic MCF‑7 cells, indicating that loss of EB1 promotes PTX‑induced stabilization of microtubules. EB1 inhibited PTX‑induced microtubule polymerization and diminished PTX binding to microtubules in vitro, suggesting that it modulates the binding sites of PTX at the growing microtubule ends. Results demonstrate that EB1 downregulates inhibition of PTX‑induced proliferation and apoptosis in breast cancer cells through a mechanism in which it impairs PTX‑mediated stabilization of microtubule polymerization and inhibits PTX binding on microtubules.
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http://dx.doi.org/10.3892/ijo.2014.2701DOI Listing
January 2015

Microtubule +TIP protein EB1 binds to GTP and undergoes dissociation from dimer to monomer on binding GTP.

Biochemistry 2014 Sep 21;53(34):5551-7. Epub 2014 Aug 21.

School of Biology, Indian Institute of Science Education and Research, Thiruvananthapuram , CET Campus, Thiruvananthapuram, Kerala 695016, India.

The +TIP protein EB1 autonomously tracks the growing plus end of microtubules and regulates plus-end dynamics. Previous studies have indicated that EB1 can recognize GTP-bound tubulin structures at the plus end, and it localizes on the microtubule surface at a site close to the exchangeable GTP-binding site of tubulin. Although the GTP-dependent structural change in tubulin has been demonstrated to be a critical determinant for recognition of plus ends by EB1, the effect of GTP on the structure of EB1 has remained unclear. Here, we have used spectroscopic, calorimetric, and biochemical methods to analyze the effect of GTP on EB1 in vitro. Isothermal titration calorimetry and tryptophan fluorescence quenching experiments demonstrated that EB1 binds to GTP with a dissociation constant ~30 μM. Circular dichroism measurements showed that EB1 undergoes changes in its secondary structure on binding GTP. Size-exclusion chromatography and urea-induced unfolding analyses revealed that GTP binding induces dissociation of the EB1 dimer to monomers. Size-exclusion chromatography followed by biochemical analysis further determined that EB1-GTP binding involves association of approximately one molecule of GTP per EB1 monomer. The results reveal a hitherto unknown GTP-dependent mechanism of dimer-to-monomer transition in EB1 and further implicate its possible role in regulating the stability of the EB1 dimer vs monomer as well as plus-end regulation in cells.
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http://dx.doi.org/10.1021/bi5007942DOI Listing
September 2014