BRCA1-BARD1: the importance of being in shape Daza-Martin, Manuel; Densham, Ruth M.; Morris, Joanna R.
Molecular & cellular oncology,
11/2019, Letnik:
6, Številka:
6
Journal Article
Odprti dostop
The breast cancer type-1 susceptibility protein (BRCA1) contributes to genome integrity through homologous recombinational DNA repair and by protecting stalled replication forks from nucleolytic ...degradation. We recently discovered that fork protection requires a conformational change of BRCA1 unimportant to homologous recombination repair, indicating separate roles for BRCA1 in these pathways.
The conditional kinase ΔMEKK3:ER⁎ allows activation of JNK, p38 and ERK1/2 without overt cellular stress or damage and has proved useful in understanding how these pathways regulate apoptosis and ...cell cycle progression. We have previously shown that activation of ΔMEKK3:ER⁎ causes a sustained G
1 cell cycle arrest which requires p21
CIP1, with ERK1/2 and p38 cooperating to promote p21
CIP1 expression. In cells lacking p21
CIP1, ΔMEKK3:ER⁎ causes only a transient delay in cell cycle re-entry. We now show that this delay in cell cycle re-entry is due to a reduction in cyclin D1 levels. Activation of ΔMEKK3:ER⁎ promotes the proteasome-dependent turnover of cyclin D1; this requires phosphorylation of threonine 286 (T
286) and expression of cyclin D1T
286A rescues the delay in G
1/S progression. ΔMEKK3:ER⁎-dependent phosphorylation of T
286 does not appear to be mediated by GSK3β but requires activation of the ERK1/2 and p38 pathways. ERK1/2 can physically associate with cyclin D1 but activation of ERK1/2 alone is not sufficient for phosphorylation of T
286. Rather, cyclin D1 phosphorylation appears to require coincident activation of ERK1/2 and p38. Thus activation of ΔMEKK3:ER⁎ promotes a sustained G
1 cell cycle arrest by a bipartite mechanism involving the rapid destruction of cyclin D1 and the slower more prolonged expression of p21
CIP1. This has parallels with the bipartite response to ionizing radiation and p53-independent mechanisms of G
1 cell cycle arrest in simple organisms such as yeast.
To study the mechanisms by which mitogen- and stress-activated protein kinases regulate cell cycle re-entry, we have used a panel of conditional kinases that stimulate defined MAPK or SAPK cascades. ...Activation of DeltaMEKK3:ER* during serum restimulation of quiescent cells causes a strong activation of JNK1 and p38alpha but only a modest potentiation of serum-stimulated ERK1/2 activity. In CCl39 cells this promoted a sustained G1 arrest that correlated with decreased expression of cyclin D1 and Cdc25A, increased expression of p21CIP1 and inhibition of CDK2 activity. In Rat-1 cells, in which p21(CIP1) expression is silenced by methylation, DeltaMEKK3:ER* activation caused only a transient delay in the S phase entry rather than a sustained G1 arrest. Furthermore, p21CIP1-/- 3T3 cells were defective for the DeltaMEKK3:ER*-induced G1 cell cycle arrest compared to their wild-type counterparts. These results suggest that activated DeltaMEKK3:ER* inhibits the G1 --> S progression by two kinetically distinct mechanisms, with expression of p21CIP1 being required to ensure a sustained G1 cell cycle arrest. The ERK1/2 and p38alphabeta pathways cooperated to induce p21CIP1 expression and inhibition of p38alphabeta caused a partial reversal of the cell cycle arrest. In contrast, selective activation of ERK1/2 by DeltaRaf-1:ER* did not inhibit serum stimulated cell cycle re-entry. Finally, selective activation of JNK by DeltaMEKK1:ER* failed to inhibit cell cycle re-entry, even in cells that retained wild-type p53, arguing against a major role for JNK alone in antagonizing the G1 --> S transition.
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