Highlights d S-CDK and Cdc5 kinases sequentially phosphorylate Sgs1 during the cell cycle d CDK phosphorylation enhances the DNA unwinding activity of Sgs1 d Phosphorylation increases the velocity and processivity of Sgs1 molecules d Phospho-stimulation of Sgs1 is required for mitotic and meiotic DNA repair

The intracellular trafficking of growth factor receptors determines the activity of their downstream signaling pathways. Here, we show that the putative HSP-90 co-chaperone CHP-1 acts as a regulator of EGFR trafficking in C. elegans. Loss of chp-1 causes the retention of the EGFR in the ER and decreases MAPK signaling. CHP-1 is specifically required for EGFR trafficking, as the localization of other transmembrane receptors is unaltered in chp-1(lf) mutants, and the inhibition of hsp-90 or other co-chaperones does not affect EGFR localization. The role of the CHP-1 homolog CHORDC1 during EGFR trafficking is conserved in human cells. Analogous to C. elegans, the response of CHORDC1-deficient A431 cells to EGF stimulation is attenuated, the EGFR accumulates in the ER and ERK2 activity decreases. Although CHP-1 has been proposed to act as a co-chaperone for HSP90, our data indicate that CHP-1 plays an HSP90-independent function in controlling EGFR trafficking through the ER.

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14The intracellular trafficking of growth factor receptors determines the activity of their 15 downstream signaling pathways. The putative co-chaperone CHP-1 acts as a regulator of EGFR 16 trafficking during C.elegans vulval development. Loss of chp-1 causes the retention of the 17 EGFR in the ER and decreased MAPK signaling. CHP-1 functions specifically, as the localization 18 of other receptors is unaltered in chp-1(lf) mutants, and inhibiting other co-chaperones does 19 not affect EGFR localization. The role of CHP-1 during EGFR trafficking is conserved in 20 humans. Analogous to C.elegans, the response of CHP-1-deficient human cells to EGF 21 stimulation is attenuated, the EGFR accumulates in the ER and ERK2 activity is decreased. 22Although CHP-1 has been proposed to act as a co-chaperone for HSP90, our data indicate an 23 HSP90-independent function of CHP-1. The identification of CHP-1 as a regulator of EGFR24 trafficking opens the possibility to identify small molecule chaperone inhibitors targeting the 25 EGFR pathway with increased selectivity. 26 Thanks to the availability of functional GFP tagged LET-23 reporters and the transparent body, vulval 55 development is an excellent model to observe EGFR trafficking and localization in the epithelial VPCs 56 of living animals (Haag et al, 2014). Before vulval induction, LET-23 is equally expressed in all VPCs. 57 During induction, a positive MAP kinase MPK-1 feedback signal up-regulates LET-23 expression in 58 P6.p, which allows this cell to sequester most of the inductive LIN-3 EGF signal (Stetak et al, 2006). 59 By contrast, LIN-12 NOTCH signaling in P5.p and P7.p results in the down-regulation of LET-23 and 60 Haag et al. 4 the inhibition of RAS/MAPK signaling (Whitfield et al, 1999). 61 Several factors that regulate RAS/MAPK activity by controlling the sub-cellular localization and 62 trafficking of the LET-23 EGFR in the VPCs have been identified. The basolateral localization of LET-63 23 by the tripartite LIN-2/CASK, LIN-10/ MINT and LIN-7/VELIS protein complex is necessary for 64 efficient receptor activation, because LIN-3 is secreted by the AC in the somatic gonad facing the 65 basolateral compartment of the VPCs (Kaech et al, 1998;Hoskins et al, 1996). On the other hand, the 66 ARF GTPase exchange factor AGEF-1 antagonizes via the ARF GTPase and the AP-1 adaptor complex 67 the basolateral localization of LET-23 (Skorobogata et al, 2014). A systematic in vivo screen in live C. 68 elegans larvae has identified multiple additional regulators of LET-23 localization and signaling 69 (Haag et al, 2014). One candidate identified in this screen was chp-1, which encodes a Cysteine and 70Histidine Rich Domain (CHORD) containing protein homologous to human CHORDC1 (also named 71Morgana) (Brancaccio et al, 2003;Ferretti et al, 2011). CHORDC1 has been proposed to function as 72 co-chaperone with HSP90, though CHORDC1 may also act independently of HPS90 (Gano & Simon, 73 2010a). 74Here, we show that a loss of chp-1 function in C. elegans leads to the accumulat...