<i>Dictyostelium</i>AMPKα regulates aggregate size and cell-type patterning

Maurya, Ranjana; Kumar, Rakesh; Saran, Shweta

doi:10.1098/rsob.170055

Cited by 16 publications

(36 citation statements)

References 58 publications

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“…Finer anatomy of wildtype and ampkα − fruiting bodies revealed alteration in stalk cell arrangements. Earlier, we showed that AMPKα gets activated upon starvation (Maurya et al, 2017) therefore we were interested to find whether deletion of ampkα rendered cell survival during starvation.…”

Section: Discussionmentioning

confidence: 99%

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AMPKα promotes basal autophagy induction in Dictyostelium discoideum

Maurya

Kumar

Saran

2019

Journal Cellular Physiology

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Autophagy is a degradation process, wherein long-lived proteins, damaged organelles, and protein aggregates are degraded to maintain cellular homeostasis. Upon starvation, 5′-AMP-activated protein kinase (AMPK) initiates autophagy. We show that ampkα − cells exhibit 50% reduction in pinocytosis and display defective phagocytosis. Re-expression of AMPKα in ampkα − cells co-localizes with red fluorescence protein-tagged bacteria. The ampkα − cells show reduced cell survival and autophagic flux under basal and starvation conditions. Co-immunoprecipitation studies show conservation of the AMPK-ATG1 axis in basal autophagy. Computational analyses suggest that the N-terminal region of DdATG1 is amenable for interaction with AMPK. Furthermore, β-actin was found to be a novel interacting partner of AMPK, attributed to the alteration in macropinocytosis and phagocytosis in the absence of AMPK. Additionally, ampkα − cells exhibit enhanced polyubiquitinated protein levels and allied large ubiquitin-positive protein aggregates.Our findings suggest that AMPK provides links among pinocytosis, phagocytosis, autophagy, and is a requisite for basal autophagy in Dictyostelium.

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Section: Discussionmentioning

confidence: 99%

“…Cells were grown in HL5 medium with or without the appropriate antibiotics (G148 or blasticidin). Dictyostelium cells were transformed by the electroporation method and ampkα null cells used in this study were prepared in our previous study (Maurya, Kumar, & Saran, 2017).…”

Section: Dictyostelium Growth Development and Transformationmentioning

confidence: 99%

AMPKα promotes basal autophagy induction in Dictyostelium discoideum

Maurya

Kumar

Saran

2019

Journal Cellular Physiology

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“…Briefly, the logarithmic phase cells (3-5x10 6 cells ml -1 ) were used for inoculating the secondary culture at a starting density of approximately 5x10 5 cells ml -1 and monitored over several days. Development analyses were performed according to Maurya et al, (2017). The logarithmic phase cells were harvested, washed in 1xKK 2 buffer and spotted at a density of 5x10 7 cells ml -1 on non-nutrient agar (NNA) plates.…”

Section: Methodsmentioning

confidence: 99%

Deletion of etoposide-induced 2.4 kb transcript (ei24) reduced cell proliferation and aggregate-size in Dictyostelium discoideum

Gupta¹,

Saran²

2018

Int. J. Dev. Biol.

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The etoposide-induced 2.4 kb transcript (ei24) gene is induced both by p53 and etoposide, an anti-cancer tumour drug. There is no p53 gene present in Dictyostelium discoideum. Thus, the functions of ei24 in the absence of p53 were analysed. Both overexpressor (ei24) and knockout (ei24) mutants were made to study its role during growth, development and differentiation. Additionally, cell cycle and its response to DNA-damage were also analysed. We identified, characterized and elucidated the functions of the ei24 gene in Dictyostelium. In silico analyses demonstrated the conservation across eukaryotes and in situ hybridization showed it to be prestalk-specific. ei24 cells showed reduced cell proliferation and cell-cohesive properties, ultimately forming small-sized aggregates that developed into miniature and stalky fruiting bodies. The ei24 cells formed fruiting bodies with engorged or double-decker type sori with short stalks. The ei24 cells showed reduced cAMP signalling with lower intracellular cAMP levels resulting in diminished migration of cells along cAMP gradients. Deletion of ei24 resulted in mis-expression of prestalk-specific markers. Cell cycle analysis revealed an increased bias towards the stalk-pathway by ei24 cells and vice-versa for ei24 cells. EI24 in Dictyostelium functions even in the absence of p53 and is induced in response to both UV-radiation and etoposide treatments. ei24 cells showed enhanced DNA-damage repair mechanisms. Also, etoposide treatment and overexpression of ei24 caused G2/M arrest in the cell cycle. Our results indicate that EI24 is important for the growth, development and differentiation of Dictyostelium apart from being a DNA-damage response gene.

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“…Cells were cutured in axenic medium HL5 at 22 o C (with or without G418) under shaken conditions according to Maurya et al, (2017) and proliferation was monitored according to Mishra et al, (2017).…”

Section: Strains and Developmentmentioning

confidence: 99%

“…Protocols for RNA isolation and RT-PCR (Reverse transcriptase-polymerase chain reaction) were followed as previously described (Maurya et al, 2017). Following cDNA synthesis, RT-PCR reactions were performed using gene-specific primer pairs ( Supplementary Table S1) for various genes.…”

Section: Treatment With Mgbg and Spermidinementioning

confidence: 99%

Overexpression of S-adenosylmethionine decarboxylase impacts polyamine homeostasis during development of Dictyostelium discoideum

Sharma

Kumar²,

Saran³

2018

Int. J. Dev. Biol.

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The polyamines putrescine, spermidine and spermine are essential polycations involved in the regulation of cellular proliferation. They exert dynamic effects on nucleic acids and macromolecular synthesis in vitro but their specific functions in vivo are poorly understood. Here, we have modulated the spermidine levels either by overexpressing the S-adenosylmethionine decarboxylase (samdc) gene or treating the cells with methylglyoxal-bis (guanylhydrazone) (MGBG), an inhibitor of SAMDC. In Dictyostelium, overexpression of SAMDC slowed cell proliferation, delayed development and arrested cells in the S-phase of the cell cycle. Treatment with MGBG reduced cell proliferation and stimulated development, but in samdc OE cells, it increased cell proliferation suggesting critical levels of spermidine to be important. In samdc OE cells, spermidine levels remained high throughout development but only small changes in the spermine levels were observed. Initial putrescine levels did increase but reverted to wild-type levels after the mound stage. As tight regulation of polyamine homeostasis is required, we identified genes that could be involved in its maintenance. In conclusion, we characterised samdc OE cells and observed the maintenance of polyamine homeostasis during the development of Dictyostelium cells.

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DictyosteliumAMPKα regulates aggregate size and cell-type patterning

Cited by 16 publications

References 58 publications

AMPKα promotes basal autophagy induction in Dictyostelium discoideum

AMPKα promotes basal autophagy induction in Dictyostelium discoideum

Deletion of etoposide-induced 2.4 kb transcript (ei24) reduced cell proliferation and aggregate-size in Dictyostelium discoideum

Overexpression of S-adenosylmethionine decarboxylase impacts polyamine homeostasis during development of Dictyostelium discoideum

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