The PTEN tumor suppressor is mutated in diverse human cancers and in hereditary cancer predisposition syndromes. PTEN is a phosphatase that can act on both polypeptide and phosphoinositide substrates in vitro. The PTEN structure reveals a phosphatase domain that is similar to protein phosphatases but has an enlarged active site important for the accommodation of the phosphoinositide substrate. The structure also reveals that PTEN has a C2 domain. The PTEN C2 domain binds phospholipid membranes in vitro, and mutation of basic residues that could mediate this reduces PTEN's membrane affinity and its ability to suppress the growth of glioblastoma tumor cells. The phosphatase and C2 domains associate across an extensive interface, suggesting that the C2 domain may serve to productively position the catalytic domain on the membrane.

Jieoh Lee    Yang Haijuan    Georgescu    Di Cristofano Antonio    Maehama Tomohiko    施一公     Jack edward Dixon    Pandolfi Pier Paolo    Nikola Pavletich   

Cell

1999

Signal transduction specificity in the transforming growth factor-β (TGF-β) system is determined by ligand activation of a receptor complex which then recruits and phosphorylates a subset of SMAD proteins including Smads 1 and 2. These then associate with Smad4 and move into the nucleus where they regulate transcription. We have identified a discrete surface structure in Smads 1 and 2 that mediates and specifies their receptor interactions. This structure is the L3 loop, a 17 amino acid region that protrudes from the core of the conserved SMAD C-terminal domain. The L3 loop sequence is invariant among TGF-β- and bone morphogenetic protein (BMP)-activated SMADS, but differs at two positions between these two groups. Swapping these two amino acids in Smads 1 and 2 induces a gain or loss, respectively, in their ability to associate with the TGF-β receptor complex and causes a switch in the phosphorylation of Smads 1 and 2 by the BMP and TGF-β receptors, respectively. A full switch in phosphorylation and activation of Smads 1 and 2 is obtained by swapping both these two amino acids and four amino acids near the C-terminal receptor phosphorylation sites. These studies identify the L3 loop as a determinant of specific SMAD-receptor interactions, and indicate that the L3 loop, together with the C-terminal tail, specifies SMAD activation.

Lo Roger S.    Chen Ye Guang    施一公     Nikola Pavletich    Joan Massagué   

EMBO Journal

1998

The Smad family of proteins, which are frequently targeted by tumorigenic mutations in cancer, mediate TGF-β signaling from cell membrane to nucleus. The crystal structure of a Smad3 MH1 domain bound to an optimal DNA sequence determined at 2.8 Å resolution reveals a novel DNA-binding motif. In the crystals, base-specific DNA recognition is provided exclusively by a conserved 11-residue β hairpin that is embedded in the major groove of DNA. A surface loop region, to which tumorigenic mutations map, has been identified as a functional surface important for Smad activity. This structure establishes a framework for understanding how Smad proteins may act in concert with other transcription factors in the regulation of TGF-β- responsive genes.

施一公     Wang Yan Fel    Jayaraman Lata    Yang Haijuan    Joan Massagué    Nikola Pavletich   

Cell

1998

Transforming growth factor-β (TGF-β) and related cytokines control the development and homeostasis of many tissues regulating the expression of genes that determine cell phenotype. Recent progress has elucidated the way in which members of the TGF-β family initiate their signal through transmembrane receptors and transmit it to target genes in via the Smad family of signal-transducing proteins. This review describes the TGF-β signaling pathways as currently understood and mutations of the genes that encode Smads that disrupt the function of these proteins and cause various forms of cancer.

Hata Phd Akiko    Joan Massagué    施一公    

Molecular Medicine Today

1998

The disassembly of the intron lariat spliceosome (ILS) marks the end of a splicing cycle. Here we report a cryoelectron microscopy structure of the ILS complex from Saccharomyces cerevisiae at an average resolution of 3.5 Å. The intron lariat remains bound in the spliceosome whereas the ligated exon is already dissociated. The step II splicing factors Prp17 and Prp18, along with Cwc21 and Cwc22 that stabilize the 5′ exon binding to loop I of U5 small nuclear RNA (snRNA), have been released from the active site assembly. The DEAH family ATPase/helicase Prp43 binds Syf1 at the periphery of the spliceosome, with its RNA-binding site close to the 3′ end of U6 snRNA. The C-terminal domain of Ntr1/Spp382 associates with the GTPase Snu114, and Ntr2 is anchored to Prp8 while interacting with the superhelical domain of Ntr1. These structural features suggest a plausible mechanism for the disassembly of the ILS complex.

Ruixue Wan;Chuangye Yan;Rui Bai;Jianlin Lei;一公 施

Cell

2017-9-21

Smad proteins are the most well-characterized intracellular effectors of the transforming growth factor β (TGF-β) signal. The ability of the Smads to act as transcriptional activators via TGF-β-induced recruitment to Smad binding elements (SBE) within the promoters of TGF-β target genes has been firmly established. However, the elucidation of the molecular mechanisms involved in TGF-β-mediated transcriptional repression are only recently being uncovered. The proto-oncogene c-myc is repressed by TGF-β, and this repression is required for the manifestation of the TGF-β cytostatic program in specific cell types. We have shown that Smad3 is required for both TGF-β-induced repression of c-myc and subsequent growth arrest in keratinocytes. The transcriptional repression of c-myc is dependent on direct Smad3 binding to a novel Smad binding site, termed a repressive Smad binding element (RSBE), within the TGF-β inhibitory element (TIE) of the c-myc promoter. The c-myc TIE is a composite element, comprised of an overlapping RSBE and a consensus E2F site, that is capable of binding at least Smad3, Smad4, E2F-4, and p107. The RSBE is distinct from the previously defined SBE and may partially dictate, in conjunction with the promoter context of the overlapping E2F site, whether the Smad3-containing complex actively represses, as opposed to transactivates, the c-myc promoter.

Joshua P. Frederick;Nicole T. Liberati;David S. Waddell;一公 施;Xiao Fan Wang

Molecular and Cellular Biology

2004-3

Presenilin is the catalytic subunit of γ-secretase, a four-component intramembrane protease responsible for the generation of β-amyloid (Aβ) peptides. Over 200 Alzheimer’s disease-related mutations have been identified in presenilin 1 (PS1) and PS2. Here, we report that Bax-inhibitor 1 (BI1), an evolutionarily conserved transmembrane protein, stably associates with PS1. BI1 specifically interacts with PS1 in isolation, but not with PS1 in the context of an assembled γ-secretase. The PS1–BI1 complex exhibits no apparent proteolytic activity, as judged by the inability to produce Aβ40 and Aβ42 from the substrate APP-C99. At an equimolar concentration, BI1 has no impact on the proteolytic activity of γ-secretase; at a 200-fold molar excess, BI1 reduces γ-secretase activity nearly by half. Our biochemical study identified BI1 as a PS1-interacting protein, suggesting additional functions of PS1 beyond its involvement in γ-secretase.

Shenjie Wu;Wenqi Song;Catherine C.L. Wong;一公 施

Proceedings of the National Academy of Sciences of the United States of America

2019-1-2

The acquisition of apoptosis resistance is a fundamental event in cancer development. Among the mechanisms used by cancer cells to evade apoptosis is the dysregulation of inhibitor of apoptosis (IAP) proteins. The activity of the IAPs is regulated by endogenous IAP antagonists such as SMAC (also termed DIABLO). Antagonism of IAP proteins by SMAC occurs via binding of the N-terminal tetrapeptide (AVPI) of SMAC to selected BIR domains of the IAPs. Small molecule compounds that mimic the AVPI motif of SMAC have been designed to overcome IAP-mediated apoptosis resistance of cancer cells. Here, we report the preclinical characterization of birinapant (TL32711), a bivalent SMAC-mimetic compound currently in clinical trials for the treatment of cancer. Birinapant bound to the BIR3 domains of cIAP1, cIAP2, XIAP, and the BIR domain of ML-IAP in vitro and induced the autoubiquitylation and proteasomal degradation of cIAP1 and cIAP2 in intact cells, which resulted in formation of a RIPK1:caspase-8 complex, caspase-8 activation, and induction of tumor cell death. Birinapant preferentially targeted the TRAF2-associated cIAP1 and cIAP2 with subsequent inhibition of TNF-induced NF-κβ activation. The activity of a variety of chemotherapeutic cancer drugs was potentiated by birinapant both in a TNF-dependent or TNF-independent manner. Tumor growth in multiple primary patient-derived xenotransplant models was inhibited by birinapant at well-tolerated doses. These results support the therapeutic combination of birinapant with multiple chemotherapies, in particular, those therapies that can induce TNF secretion. Mol Cancer Ther; 13(4); 867-79.

Christopher A. Benetatos;Yasuhiro Mitsuuchi;Jennifer M. Burns;Eric M. Neiman;Stephen M. Condon;Guangyao Yu;Martin E. Seipel;Gurpreet S. Kapoor;Matthew G. LaPorte;Susan R. Rippin;Yijun Deng;Mukta S. Hendi;Pavan K. Tirunahari;Yu Hua Lee;Thomas Haimowitz;Matthew D. Alexander;Martin A. Graham;David Weng;一公 施;Mark A. McKinlay;Srinivas K. Chunduru

Molecular Cancer Therapeutics

2014-4

Zinc finger domains of the Cys₂His₂ type are found in a large number of eukaryotic proteins. Various proteins containing these domains have been shown to bind specifically to DNA, RNA, and DNA-RNA hybrids. Structural studies of zinc finger protein-DNA complexes have revealed that the DNA molecules are underwound relative to canonical B-form. It has not been clear if zinc finger proteins recognize preexisting underwound conformations of DNA or if they induce such conformations upon binding. We report that the DNA binding domains of Sp1 and several designed zinc finger proteins unwind DNA upon binding. The extent of unwinding is consistent with that observed in zinc finger protein-DNA cocrystal structures. These DNA deformations may be important in determining overall binding affinities as well as influencing binding site preferences. Furthermore, changes in DNA conformation upon zinc finger protein binding may affect protein-protein interactions important for transcriptional regulation and other activities of zinc finger proteins.

一公 施;Jeremy M. Berg

Biochemistry

1996-3-26

Virulent enteric pathogens such as Escherichia coli strain O157:H7 rely on acid-resistance (AR) systems to survive the acidic environment in the stomach. A major component of AR is an arginine-dependent arginine:agmatine antiporter that expels intracellular protons. Here, we report the crystal structure of AdiC, the arginine:agmatine antiporter from E. coli O157:H7 and a member of the amino acid/polyamine/organocation (APC) superfamily of transporters at 3.6 Å resolution. The overall fold is similar to that of several Na ⁺-coupled symporters. AdiC contains 12 transmembrane segments, forms a homodimer, and exists in an outward-facing, open conformation in the crystals. A conserved, acidic pocket opens to the periplasm. Structural and biochemical analysis reveals the essential ligand-binding residues, defines the transport route, and suggests a conserved mechanism for the antiporter activity.

Xiang Gao;Feiran Lu;Lijun Zhou;Shangyu Dang;Linfeng Sun;Xiaochun Li;Jiawei Wang;一公 施

Science

2009-6-19