Cholesterol(CHL)serves as a building block for membrane biogenesis and a precursor to oxysterols,steroid hormones,bile acids,and vitamin D.The lysosome serves as a major sorting station for low-density lipoproteins(LD...Cholesterol(CHL)serves as a building block for membrane biogenesis and a precursor to oxysterols,steroid hormones,bile acids,and vitamin D.The lysosome serves as a major sorting station for low-density lipoproteins(LDLs),which carry dietary CHL,and it is also the cellular site where the master growth regulator,the protein kinase mechanistic Target of Rapamycin Complex 1(mTORC1),is activated.Recently,the lysosomal transmembrane protein GPR155 was reported to signals CHL sufficiency to mTORC1 through sequestration of the GTPase-activating protein towards the Rags 1(GATOR1).Although the recently reported structures of GPR155 have revealed the CHL binding site,how the signal is transduced from the CHL binding site to the soluble parts of GPR155 and GATOR1 remains unknown.Here,with our three cryo-EM structures of GPR155 captured in different conformations in complex with CHL,complemented by long-time scale molecular dynamics simulations,the dynamic rearrangement of different domains was observed.CHL binding induces a widening of the crevice between the transporter and GPCR domains.The extending helix preceding transmembrane helix(TM)16,which was unresolved in other structures,acts as a linkage lever that transmits the rotation of the GPCR domain to the soluble parts of GPR155 in response to CHL binding.This work not only answers the question of how CHL is sensed by GPR155,but also addresses a more profound question:how the signal perceived by the TMs regions is transduced to the LED and DEP domains.展开更多
Focal epilepsy accounts for 60% of all forms of epilepsy, but the pathogenic mechanism is not well understood. In this study,three novel mutations in NPRL3(nitrogen permease regulator-like 3), c.937_945del, c.1514dup ...Focal epilepsy accounts for 60% of all forms of epilepsy, but the pathogenic mechanism is not well understood. In this study,three novel mutations in NPRL3(nitrogen permease regulator-like 3), c.937_945del, c.1514dup C and 6,706-bp genomic DNA(g DNA) deletion, were identified in three families with focal epilepsy by linkage analysis, whole exome sequencing(WES) and Sanger sequencing. NPRL3 protein is a component of the GATOR1 complex, a major inhibitor of m TOR signaling. These mutations led to truncation of the NPRL3 protein and hampered the binding between NPRL3 and DEPDC5, which is another component of the GATOR1 complex. Consequently, the mutant proteins enhanced m TOR signaling in cultured cells, possibly due to impaired inhibition of m TORC1 by GATOR1. Knockdown of nprl3 in Drosophila resulted in epilepsy-like behavior and abnormal synaptic development. Taken together, these findings expand the genotypic spectrum of NPRL3-associated focal epilepsy and provide further insight into how NPRL3 mutations lead to epilepsy.展开更多
基金supported by the Scientific and Technological Innovation 2030(2022ZD0211905)the National Natural Science Foundation of China(NSFC-SNSF 32161133022)+1 种基金the Fundamental Research Program of Shanxi Province(202403021211192)the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-KJGG-022 and TSBICIP-IJCP-001).
文摘Cholesterol(CHL)serves as a building block for membrane biogenesis and a precursor to oxysterols,steroid hormones,bile acids,and vitamin D.The lysosome serves as a major sorting station for low-density lipoproteins(LDLs),which carry dietary CHL,and it is also the cellular site where the master growth regulator,the protein kinase mechanistic Target of Rapamycin Complex 1(mTORC1),is activated.Recently,the lysosomal transmembrane protein GPR155 was reported to signals CHL sufficiency to mTORC1 through sequestration of the GTPase-activating protein towards the Rags 1(GATOR1).Although the recently reported structures of GPR155 have revealed the CHL binding site,how the signal is transduced from the CHL binding site to the soluble parts of GPR155 and GATOR1 remains unknown.Here,with our three cryo-EM structures of GPR155 captured in different conformations in complex with CHL,complemented by long-time scale molecular dynamics simulations,the dynamic rearrangement of different domains was observed.CHL binding induces a widening of the crevice between the transporter and GPCR domains.The extending helix preceding transmembrane helix(TM)16,which was unresolved in other structures,acts as a linkage lever that transmits the rotation of the GPCR domain to the soluble parts of GPR155 in response to CHL binding.This work not only answers the question of how CHL is sensed by GPR155,but also addresses a more profound question:how the signal perceived by the TMs regions is transduced to the LED and DEP domains.
基金supported by the National Natural Science Foundation of China (32270663, 31871262, U20A20355,32022035)Shanghai Municipal Science and Technology Major Project(2018SHZDZX05)the Ministry of Science and Technology of China STI2030-Major Projects (2021ZD0203202)。
文摘Focal epilepsy accounts for 60% of all forms of epilepsy, but the pathogenic mechanism is not well understood. In this study,three novel mutations in NPRL3(nitrogen permease regulator-like 3), c.937_945del, c.1514dup C and 6,706-bp genomic DNA(g DNA) deletion, were identified in three families with focal epilepsy by linkage analysis, whole exome sequencing(WES) and Sanger sequencing. NPRL3 protein is a component of the GATOR1 complex, a major inhibitor of m TOR signaling. These mutations led to truncation of the NPRL3 protein and hampered the binding between NPRL3 and DEPDC5, which is another component of the GATOR1 complex. Consequently, the mutant proteins enhanced m TOR signaling in cultured cells, possibly due to impaired inhibition of m TORC1 by GATOR1. Knockdown of nprl3 in Drosophila resulted in epilepsy-like behavior and abnormal synaptic development. Taken together, these findings expand the genotypic spectrum of NPRL3-associated focal epilepsy and provide further insight into how NPRL3 mutations lead to epilepsy.
