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Abstract The transcriptional repressor DREAM (downstream regulatory element antagonist modulator) is a multifunctional neuronal calcium sensor (NCS) that controls Ca2+ and protein homeostasis through gene regulation and protein-protein interactions. Downregulation of DREAM is part of an endogenous neuroprotective mechanism that improves ATF6 (activating transcription factor 6) processing, neuronal survival in the striatum, and motor coordination in R6/2 mice, a model of Huntington's disease (HD). Whether modulation of DREAM activity can also ameliorate cognition deficits in HD mice has not been studied. Moreover, it is not known whether DREAM downregulation in HD is unique, or also occurs for other NCS family members. Using the novel object recognition test, we show that chronic administration of the DREAM-binding molecule repaglinide, or induced DREAM haplodeficiency delays onset of cognitive impairment in R6/1 mice, another HD model. The mechanism involves a notable rise in the levels of transcriptionally active ATF6 protein in the hippocampus after repaglinide administration. In addition, we show that reduction in DREAM protein in the hippocampus of HD patients was not accompanied by downregulation of other NCS family members. Our results indicate that DREAM inhibition markedly improves ATF6 processing in the hippocampus and that it might contribute to a delay in memory decline in HD mice. The mechanism of neuroprotection through DREAM silencing in HD does not apply to other NCS family members. ; This work was funded by the Instituto de Salud Carlos III/CIBERNED (to JRN, BM and AR), the Madrid regional government/Neurodegmodels (to JRN), and SAF2014–53412-R and SAF2017–89554-R (AEI-FEDER, EU) (to JRN).

DREAM, a neuronal calcium sensor protein, has multiple cellular roles including the regulation of Ca2+ and protein homeostasis. We recently showed that reduced DREAM expression or blockade of DREAM activity by repaglinide is neuroprotective in Huntington's disease (HD). Here we used structure-based drug design to guide the identification of IQM-PC330, which was more potent and had longer lasting effects than repaglinide to inhibit DREAM in cellular and in vivo HD models. We disclosed and validated an unexplored ligand binding site, showing Tyr118 and Tyr130 as critical residues for binding and modulation of DREAM activity. IQM-PC330 binding de-repressed c-fos gene expression, silenced the DREAM effect on KV4.3 channel gating and blocked the ATF6/DREAM interaction. Our results validate DREAM as a valuable target and propose more effective molecules for HD treatment ; This work was funded by the Spanish Ministery of Economy, Industry and Competitivity (AEI-FEDER, EU grants): BFU2015-67284-R (to M.G.R., R.H.), SAF2017-89554-R (to J.R.N.), SAF2016-75021-R (to C.V.) and SAF2015-66275-C2-2-R (to M.M.M.); the Instituto de Salud Carlos III CIBERNED and CIBERCV programs (to J.R.N. and to C.V., respectively) and the Madrid regional government/Neurodegmodels (to J.R.N.); CSIC grant PIE201880E109 (to M.G.R., R.H. and M.M.M.) and PIE201820E104 (to C.V.)

DREAM, a neuronal calcium sensor protein, has multiple cellular roles including the regulation of Ca2+ and protein homeostasis. We recently showed that reduced DREAM expression or blockade of DREAM activity by repaglinide is neuroprotective in Huntington's disease (HD). Here we used structure-based drug design to guide the identification of IQM-PC330, which was more potent and had longer lasting effects than repaglinide to inhibit DREAM in cellular and in vivo HD models. We disclosed and validated an unexplored ligand binding site, showing Tyr118 and Tyr130 as critical residues for binding and modulation of DREAM activity. IQM-PC330 binding de-repressed c-fos gene expression, silenced the DREAM effect on KV4.3 channel gating and blocked the ATF6/DREAM interaction. Our results validate DREAM as a valuable target and propose more effective molecules for HD treatment. ; PC was the recipient of a postgraduate FPI fellowship from the Spanish Ministry de Economy, Industry and Competitivity (MINECO). P.C. also thanks the Spanish Society of Therapeutic Chemistry (SEQT) for a Jassen-Cilag Award for young researchers (XVIII Edition, 2016). T.G. holds a Ramón y Cajal contract. D.A.P. holds a CSIC contract. This work was funded by the Spanish Ministery of Economy, Industry and Competitivity (AEI-FEDER, EU grants): BFU2015-67284-R (to M.G.R., R.H.), SAF2017-89554-R (to J.R.N.), SAF2016-75021-R (to C.V.) and SAF2015-66275-C2-2-R (to M.M.M.); the Instituto de Salud Carlos III CIBERNED and CIBERCV programs (to J.R.N. and to C.V., respectively) and the Madrid regional government/Neurodegmodels (to J.R.N.); CSIC grant PIE201880E109 (to M.G.R., R.H. and M.M.M.) and PIE201820E104 (to C.V.). ; Peer reviewed

Deregulated protein and Ca2+ homeostasis underlie synaptic dysfunction and neurodegeneration in Huntington disease (HD); however, the factors that disrupt homeostasis are not fully understood. Here, we determined that expression of downstream regulatory element antagonist modulator (DREAM), a multifunctional Ca2+-binding protein, is reduced in murine in vivo and in vitro HD models and in HD patients. DREAM downregulation was observed early after birth and was associated with endogenous neuroprotection. In the R6/2 mouse HD model, induced DREAM haplodeficiency or blockade of DREAM activity by chronic administration of the drug repaglinide delayed onset of motor dysfunction, reduced striatal atrophy, and prolonged life span. DREAM-related neuroprotection was linked to an interaction between DREAM and the unfolded protein response (UPR) sensor activating transcription factor 6 (ATF6). Repaglinide blocked this interaction and enhanced ATF6 processing and nuclear accumulation of transcriptionally active ATF6, improving prosurvival UPR function in striatal neurons. Together, our results identify a role for DREAM silencing in the activation of ATF6 signaling, which promotes early neuroprotection in HD. ; A. De la Cruz holds a RECAVA contract, A. Prieto and P. Cercós hold FPI fellowships, and T. González holds a Ramón y Cajal contract. J. Casado-Vela holds a JAE-DOC (CSIC) from the Spanish Ministerio de Economía y Competitividad (MINECO), cofunded by the European Social Fund. This work was funded by the Instituto de Salud Carlos III/CIBERNED (to J.R. Naranjo, B. Mellström, and A. Rábano), FISS-RIC RD12/0042/0019 (to C. Valenzuela), Madrid regional government/Neurodegmodels (to J.R. Naranjo), MINECO grants SAF2010-21784 and SAF2014-53412-R (to J.R. Naranjo), SAF2012-32209 (to M. Gutierrez-Rodriguez), SAF2010-14916 and SAF2013-45800-R (to C. Valenzuela), and a grant from the Swedish Research Council (J.Y. Li). ; Peer Reviewed

Deregulated protein and Ca2+ homeostasis underlie synaptic dysfunction and neurodegeneration in Huntington disease (HD); however, the factors that disrupt homeostasis are not fully understood. Here, we determined that expression of downstream regulatory element antagonist modulator (DREAM), a multifunctional Ca2+-binding protein, is reduced in murine in vivo and in vitro HD models and in HD patients. DREAM downregulation was observed early after birth and was associated with endogenous neuroprotection. In the R6/2 mouse HD model, induced DREAM haplodeficiency or blockade of DREAM activity by chronic administration of the drug repaglinide delayed onset of motor dysfunction, reduced striatal atrophy, and prolonged life span. DREAM-related neuroprotection was linked to an interaction between DREAM and the unfolded protein response (UPR) sensor activating transcription factor 6 (ATF6). Repaglinide blocked this interaction and enhanced ATF6 processing and nuclear accumulation of transcriptionally active ATF6, improving prosurvival UPR function in striatal neurons. Together, our results identify a role for DREAM silencing in the activation of ATF6 signaling, which promotes early neuroprotection in HD ; This work was funded by the Instituto de Salud Carlos III/CIBERNED (to J.R. Naranjo, B. Mellström, and A. Rábano), FISS-RIC RD12/0042/0019 (to C. Valenzuela), Madrid regional government/Neurodegmodels (to J.R. Naranjo), MINECO grants SAF2010-21784 and SAF2014-53412-R (to J.R. Naranjo), SAF2012-32209 (to M. Gutierrez-Rodriguez), SAF2010-14916 and SAF2013-45800-R (to C. Valenzuela), and a grant from the Swedish Research Council (J.Y. Li)