Limits...
Essential and Checkpoint Functions of Budding Yeast ATM and ATR during Meiotic Prophase Are Facilitated by Differential Phosphorylation of a Meiotic Adaptor Protein, Hop1.

Penedos A, Johnson AL, Strong E, Goldman AS, Carballo JA, Cha RS - PLoS ONE (2015)

Bottom Line: A hallmark of the conserved ATM/ATR signalling is its ability to mediate a wide range of functions utilizing only a limited number of adaptors and effector kinases.In the absence of Dmc1, the phospho-S298 also promotes Mek1 hyper-activation necessary for implementing meiotic checkpoint arrest.Taking these observations together, we propose that the Hop1 phospho-T318 and phospho-S298 constitute key components of the Tel1/Mec1- based meiotic recombination surveillance (MRS) network and facilitate effective coupling of meiotic recombination and progression during both unperturbed and challenged meiosis.

View Article: PubMed Central - PubMed

Affiliation: Division of Stem Cell Biology and Developmental Genetics, MRC National Institute for Medical Research, London, NW7 1AA, United Kingdom.

ABSTRACT
A hallmark of the conserved ATM/ATR signalling is its ability to mediate a wide range of functions utilizing only a limited number of adaptors and effector kinases. During meiosis, Tel1 and Mec1, the budding yeast ATM and ATR, respectively, rely on a meiotic adaptor protein Hop1, a 53BP1/Rad9 functional analog, and its associated kinase Mek1, a CHK2/Rad53-paralog, to mediate multiple functions: control of the formation and repair of programmed meiotic DNA double strand breaks, enforcement of inter-homolog bias, regulation of meiotic progression, and implementation of checkpoint responses. Here, we present evidence that the multi-functionality of the Tel1/Mec1-to-Hop1/Mek1 signalling depends on stepwise activation of Mek1 that is mediated by Tel1/Mec1 phosphorylation of two specific residues within Hop1: phosphorylation at the threonine 318 (T318) ensures the transient basal level Mek1 activation required for viable spore formation during unperturbed meiosis. Phosphorylation at the serine 298 (S298) promotes stable Hop1-Mek1 interaction on chromosomes following the initial phospho-T318 mediated Mek1 recruitment. In the absence of Dmc1, the phospho-S298 also promotes Mek1 hyper-activation necessary for implementing meiotic checkpoint arrest. Taking these observations together, we propose that the Hop1 phospho-T318 and phospho-S298 constitute key components of the Tel1/Mec1- based meiotic recombination surveillance (MRS) network and facilitate effective coupling of meiotic recombination and progression during both unperturbed and challenged meiosis.

No MeSH data available.


Related in: MedlinePlus

Hop1-S298 phosphorylation promotes stable Mek1-Hop1 interaction on chromosomes.(A) Hop1 and Mek1-HA chromosome association during dmc1Δ meiosis at 23°C in a HOP1, hop1-S298A, or hop1-T318A background. (B) and (C) Effects of hop1-S298A or hop1-T318A on Hop1 chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Hop1 foci was scored. (D) and (E) Effects of hop1-S298A or hop1-T318A on Mek1-HA chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Mek1-HA foci was scored. (F) and (G) Effects of hop1-S298A and hop1-T318A on Hop1-Mek1 co-localization during DMC1 and dmc1Δ meiosis. Fraction of nuclei where more than 80% of Mek1-HA foci co-localized with Hop1 foci was scored. (B-G) Errors were calculated from the 95% confidence interval of a binomial distribution. (H) and (I) Effects of hop1-S298A on Hop1-Mek1 interaction on chromosomes. Nuclear spreads of HOP1 dmc1Δ and hop1-S298A dmc1Δ were prepared from samples taken at 6 hours after induction of synchronous meiosis at 23°C. The spreads were stained with DAPI and the antibodies against Hop1 and HA (for detection of Mek1-HA).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4520594&req=5

pone.0134297.g004: Hop1-S298 phosphorylation promotes stable Mek1-Hop1 interaction on chromosomes.(A) Hop1 and Mek1-HA chromosome association during dmc1Δ meiosis at 23°C in a HOP1, hop1-S298A, or hop1-T318A background. (B) and (C) Effects of hop1-S298A or hop1-T318A on Hop1 chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Hop1 foci was scored. (D) and (E) Effects of hop1-S298A or hop1-T318A on Mek1-HA chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Mek1-HA foci was scored. (F) and (G) Effects of hop1-S298A and hop1-T318A on Hop1-Mek1 co-localization during DMC1 and dmc1Δ meiosis. Fraction of nuclei where more than 80% of Mek1-HA foci co-localized with Hop1 foci was scored. (B-G) Errors were calculated from the 95% confidence interval of a binomial distribution. (H) and (I) Effects of hop1-S298A on Hop1-Mek1 interaction on chromosomes. Nuclear spreads of HOP1 dmc1Δ and hop1-S298A dmc1Δ were prepared from samples taken at 6 hours after induction of synchronous meiosis at 23°C. The spreads were stained with DAPI and the antibodies against Hop1 and HA (for detection of Mek1-HA).

Mentions: Next, we assessed the effects of hop1-S298A on Hop1- and Mek1- chromosome association. In a DMC1 background, hop1-T318A cells exhibited a modest reduction in transient Hop1-chromosome association and no detectable signal in Mek1 association (Fig 4B and 4D). In a hop1-S298A DMC1 background, both Hop1- and Mek1-chromosome association occurred normally (Fig 4B and 4D), supporting the observation above that the phospho-S298 is dispensable for the essential Mek1 activation during normal meiosis. In the absence of DMC1, the dmc1Δ-dependent maintenance of Hop1/Mek1 chromosome association was impaired in a hop1-S298A as well as a hop1-T319A background (Fig 4A, 4C and 4D).


Essential and Checkpoint Functions of Budding Yeast ATM and ATR during Meiotic Prophase Are Facilitated by Differential Phosphorylation of a Meiotic Adaptor Protein, Hop1.

Penedos A, Johnson AL, Strong E, Goldman AS, Carballo JA, Cha RS - PLoS ONE (2015)

Hop1-S298 phosphorylation promotes stable Mek1-Hop1 interaction on chromosomes.(A) Hop1 and Mek1-HA chromosome association during dmc1Δ meiosis at 23°C in a HOP1, hop1-S298A, or hop1-T318A background. (B) and (C) Effects of hop1-S298A or hop1-T318A on Hop1 chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Hop1 foci was scored. (D) and (E) Effects of hop1-S298A or hop1-T318A on Mek1-HA chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Mek1-HA foci was scored. (F) and (G) Effects of hop1-S298A and hop1-T318A on Hop1-Mek1 co-localization during DMC1 and dmc1Δ meiosis. Fraction of nuclei where more than 80% of Mek1-HA foci co-localized with Hop1 foci was scored. (B-G) Errors were calculated from the 95% confidence interval of a binomial distribution. (H) and (I) Effects of hop1-S298A on Hop1-Mek1 interaction on chromosomes. Nuclear spreads of HOP1 dmc1Δ and hop1-S298A dmc1Δ were prepared from samples taken at 6 hours after induction of synchronous meiosis at 23°C. The spreads were stained with DAPI and the antibodies against Hop1 and HA (for detection of Mek1-HA).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4520594&req=5

pone.0134297.g004: Hop1-S298 phosphorylation promotes stable Mek1-Hop1 interaction on chromosomes.(A) Hop1 and Mek1-HA chromosome association during dmc1Δ meiosis at 23°C in a HOP1, hop1-S298A, or hop1-T318A background. (B) and (C) Effects of hop1-S298A or hop1-T318A on Hop1 chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Hop1 foci was scored. (D) and (E) Effects of hop1-S298A or hop1-T318A on Mek1-HA chromosome association during DMC1 or dmc1Δ meiosis. Fraction of cells exhibiting 10 or more Mek1-HA foci was scored. (F) and (G) Effects of hop1-S298A and hop1-T318A on Hop1-Mek1 co-localization during DMC1 and dmc1Δ meiosis. Fraction of nuclei where more than 80% of Mek1-HA foci co-localized with Hop1 foci was scored. (B-G) Errors were calculated from the 95% confidence interval of a binomial distribution. (H) and (I) Effects of hop1-S298A on Hop1-Mek1 interaction on chromosomes. Nuclear spreads of HOP1 dmc1Δ and hop1-S298A dmc1Δ were prepared from samples taken at 6 hours after induction of synchronous meiosis at 23°C. The spreads were stained with DAPI and the antibodies against Hop1 and HA (for detection of Mek1-HA).
Mentions: Next, we assessed the effects of hop1-S298A on Hop1- and Mek1- chromosome association. In a DMC1 background, hop1-T318A cells exhibited a modest reduction in transient Hop1-chromosome association and no detectable signal in Mek1 association (Fig 4B and 4D). In a hop1-S298A DMC1 background, both Hop1- and Mek1-chromosome association occurred normally (Fig 4B and 4D), supporting the observation above that the phospho-S298 is dispensable for the essential Mek1 activation during normal meiosis. In the absence of DMC1, the dmc1Δ-dependent maintenance of Hop1/Mek1 chromosome association was impaired in a hop1-S298A as well as a hop1-T319A background (Fig 4A, 4C and 4D).

Bottom Line: A hallmark of the conserved ATM/ATR signalling is its ability to mediate a wide range of functions utilizing only a limited number of adaptors and effector kinases.In the absence of Dmc1, the phospho-S298 also promotes Mek1 hyper-activation necessary for implementing meiotic checkpoint arrest.Taking these observations together, we propose that the Hop1 phospho-T318 and phospho-S298 constitute key components of the Tel1/Mec1- based meiotic recombination surveillance (MRS) network and facilitate effective coupling of meiotic recombination and progression during both unperturbed and challenged meiosis.

View Article: PubMed Central - PubMed

Affiliation: Division of Stem Cell Biology and Developmental Genetics, MRC National Institute for Medical Research, London, NW7 1AA, United Kingdom.

ABSTRACT
A hallmark of the conserved ATM/ATR signalling is its ability to mediate a wide range of functions utilizing only a limited number of adaptors and effector kinases. During meiosis, Tel1 and Mec1, the budding yeast ATM and ATR, respectively, rely on a meiotic adaptor protein Hop1, a 53BP1/Rad9 functional analog, and its associated kinase Mek1, a CHK2/Rad53-paralog, to mediate multiple functions: control of the formation and repair of programmed meiotic DNA double strand breaks, enforcement of inter-homolog bias, regulation of meiotic progression, and implementation of checkpoint responses. Here, we present evidence that the multi-functionality of the Tel1/Mec1-to-Hop1/Mek1 signalling depends on stepwise activation of Mek1 that is mediated by Tel1/Mec1 phosphorylation of two specific residues within Hop1: phosphorylation at the threonine 318 (T318) ensures the transient basal level Mek1 activation required for viable spore formation during unperturbed meiosis. Phosphorylation at the serine 298 (S298) promotes stable Hop1-Mek1 interaction on chromosomes following the initial phospho-T318 mediated Mek1 recruitment. In the absence of Dmc1, the phospho-S298 also promotes Mek1 hyper-activation necessary for implementing meiotic checkpoint arrest. Taking these observations together, we propose that the Hop1 phospho-T318 and phospho-S298 constitute key components of the Tel1/Mec1- based meiotic recombination surveillance (MRS) network and facilitate effective coupling of meiotic recombination and progression during both unperturbed and challenged meiosis.

No MeSH data available.


Related in: MedlinePlus