Nuclear export inhibition through covalent conjugation and hydrolysis of Leptomycin B by CRM1

Sun, Qingxiang; Carrasco, Yazmin P.; Hu, Youcai; Guo, Xiaofeng; Mirzaei, Hamid; MacMillan, John B.; Chook, Yuh Min

doi:10.1073/pnas.1217203110

Cited by 179 publications

(250 citation statements)

References 21 publications

(32 reference statements)

Supporting

Mentioning

242

Contrasting

Unclassified

Order By: Relevance

“…An analysis of available structural data shows that K142 R is positioned toward the Importin-β and Crm1 Huntington, elongation factor 3, PR65/A, TOR (HEAT) repeat region (Table S2; PDB ID codes 1IBR, 2HB2, 3GJX, 3NC1, and 3NBY) (3,12,49,50). Acetylation at this position might therefore interfere with import/ export receptor binding.…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, K152 R and K37 R form direct salt bridges toward the Crm1 D436, located in the Crm1 intra-HEAT9 loop known to affect export substrate release (3,49,52). K152 R and K37 R also both intramolecularly contact the acidic Ran C-terminal 211 DEDDDL 216 motif in the ternary complexes of Ran and RanGAP, as well as Ran, Crm1, and RanBP1 (Table S2; PDB ID codes 1K5D, 1K5G, and 4HAT) (50,53). Therefore, acetylation might play a role in RanGAP-catalyzed nucleotide hydrolysis and export substrate release in the presence of RanBP1.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Small GTP-binding protein Ran is regulated by posttranslational lysine acetylation

Boor

Knyphausen

Kuhlmann

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Ran is a small GTP-binding protein of the Ras superfamily regulating fundamental cellular processes: nucleo-cytoplasmic transport, nuclear envelope formation and mitotic spindle assembly. An intracellular Ran•GTP/Ran•GDP gradient created by the distinct subcellular localization of its regulators RCC1 and RanGAP mediates many of its cellular effects. Recent proteomic screens identified five Ran lysine acetylation sites in human and eleven sites in mouse/rat tissues. Some of these sites are located in functionally highly important regions such as switch I and switch II. Here, we show that lysine acetylation interferes with essential aspects of Ran function: nucleotide exchange and hydrolysis, subcellular Ran localization, GTP hydrolysis, and the interaction with import and export receptors. Deacetylation activity of certain sirtuins was detected for two Ran acetylation sites in vitro. Moreover, Ran was acetylated by CBP/p300 and Tip60 in vitro and on transferase overexpression in vivo. Overall, this study addresses many important challenges of the acetylome field, which will be discussed.Ran | lysine acetylation | genetic code expansion concept | nucleus | nuclear cytosolic transport T he small GTP-binding protein Ran (Ras-related nuclear) is involved in nucleo-cytoplasmic transport processes, nuclear envelope formation, and the formation of the mitotic spindle (1). Ran, furthermore, has a variety of cytosolic functions and is involved in the cross-talk with the actin cytoskeleton. As a member of the Ras superfamily, Ran is structurally composed of a fold known as the G-domain (GTP-binding domain), a central sixstranded β-sheet that is surrounded by α-helices. Ras-family members bind to GTP and GDP nucleotides with high picomolar affinity. However, only in the GTP-bound form and the switch Iand switch II-loops adopt a stable conformation. Ran has been structurally characterized in great detail, including different nucleotide states and various protein complexes (2-4).In interphase cells, about 90% of cellular Ran is nuclear, and only a minor proportion is cytosolic (5). The localization of the guanine-nucleotide exchange factor (GEF) RCC1 (Regulator of chromosome condensation 1) at the nuclear chromatin and the RanGAP (RanGTPase-activating protein) at the cytosolic site of the nuclear pore creates a gradient of Ran•GTP in the nucleus and Ran•GDP in the cytosol (6-8).In the nucleus, Ran•GTP binds to exportins such as CRM1 (Chromosome region maintenance 1) to transport cargo proteins containing a nuclear export signal (NES) into the cytosol (3, 9, 10). Ran•GTP, furthermore, binds to Importin-β•cargo complexes to release the cargo in the nucleus (11-15). In the cytosol, the Importin•Ran•GTP complexes, as well as the ternary exportin•Ran•GTP-cargo complexes, dissociate on binding of RanBP1 and subsequent GTP hydrolysis catalyzed by RanGAP (16,17). The Ran transport cycle closes by translocation of Ran•GDP to the nucleus by the nuclear transport factor 2 (NTF2) (4,(17)(18)(19)(20). Many of these Ran interactions also ...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Small GTP-binding protein Ran is regulated by posttranslational lysine acetylation

Boor

Knyphausen

Kuhlmann

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…It is known that leptomycin B covalently modifies chromosomal region maintenance 1 (CRM1; exportin 1) at the nucleophillic sulphydryl group of Cys 528 by utilizing its α,β‐unsaturated δ‐lactone, thus preventing export of protein cargoes that rely on this cleft by preventing formation of the ternary CRM1/cargo substrate/Ran complex, or the binary complex CRM1/cargo substrate in the absence of Ran 40. More recent computational studies of inhibitors, such as leptomycin B, to CRM1 revealed that the mechanism of inhibition goes beyond simple Michael addition, and is followed by a CRM1‐mediated hydrolysis of the lactone 41. Mutagenesis revealed that at least one of the residues Arg 543, Lys 548, or Cys 579 needs to be present to stabilize the resulting anionic tetrahedral intermediate and lower the energy barrier to drive the reaction.…”

Section: Natural Product Derived Fragments In Drug Discoverymentioning

confidence: 99%

Capturing Biological Activity in Natural Product Fragments by Chemical Synthesis

2016

View full text Add to dashboard Cite

Natural products have had an immense influence on science and have directly led to the introduction of many drugs. Organic chemistry, and its unique ability to tailor natural products through synthesis, provides an extraordinary approach to unlock the full potential of natural products. In this Review, an approach based on natural product derived fragments is presented that can successfully address some of the current challenges in drug discovery. These fragments often display significantly reduced molecular weights, reduced structural complexity, a reduced number of synthetic steps, while retaining or even improving key biological parameters such as potency or selectivity. Examples from various stages of the drug development process up to the clinic are presented. In addition, this process can be leveraged by recent developments such as genome mining, antibody–drug conjugates, and computational approaches. All these concepts have the potential to identify the next generation of drug candidates inspired by natural products.

show abstract

“…25 Based on these findings, we investigated the ability of CRM1 mutation to abrogate S109 activity. We transiently transfected HCT-15 cells with wild type and Cys528 mutant CRM1 and tested the effect of S109 on RanBP1 nuclear export.…”

Section: S109 Decreases Crm1 Protein Expression Via Proteasomalmediatmentioning

confidence: 99%

“…25 To investigate whether S109 reversibly binds to CRM1, we analyzed the subcellular localization of RanBP1 after the removal of S109-containing medium. As shown in Fig.…”

Section: S109 Decreases Crm1 Protein Expression Via Proteasomalmediatmentioning

confidence: 99%

Novel reversible selective inhibitor of nuclear export shows that CRM1 is a target in colorectal cancer cells

Niu

Chong

Han

et al. 2015

Cancer Biology & Therapy

View full text Add to dashboard Cite

Abbreviations: CRM1, chromosomal region maintenance 1; DMSO, dimethyl sulfoxide; EGFR, epidermal growth factor receptor; LMB, leptomycin B; NES, leucine-rich nuclear export signal; PI3K, phosphoinositide 3-kinase; RanBP1, Ran-binding protein 1.Colorectal cancer arises via a multistep carcinogenic process and the deregulation of multiple pathways. Thus, the simultaneous targeting of multiple pathways may be a promising therapeutic approach for colorectal treatment. CRM1 is an attractive cancer drug target, because it can regulate multiple pathways and tumor suppressor proteins. In this study, we investigated the anti-tumor activity of a novel reversible CRM1 inhibitor S109 in colorectal cancer. Our data demonstrate that S109 inhibits proliferation and induces cell cycle arrest in colorectal cancer cells. Mechanistically, we demonstrate that the activity of S109 is associated with the nuclear retention of major tumor suppress proteins. Furthermore, the Cys528 mutation of CRM1 prevented the ability of S109 to block nuclear export and inhibit the proliferation of colorectal cancer cells. Interestingly, S109 decreased the CRM1 protein level via proteasomal pathway. We argue that reversible CRM1 inhibitors but not irreversible inhibitors can induce the degradation of CRM1, because the dissociation of reversible inhibitors of CRM1 changes the conformation of CRM1. Taken together, these findings demonstrate that CRM1 is a valid target for the treatment of colorectal cancer and provide a basis for the development of S109 therapies for colorectal cancer.

show abstract

Nuclear export inhibition through covalent conjugation and hydrolysis of Leptomycin B by CRM1

Cited by 179 publications

References 21 publications

Small GTP-binding protein Ran is regulated by posttranslational lysine acetylation

Small GTP-binding protein Ran is regulated by posttranslational lysine acetylation

Capturing Biological Activity in Natural Product Fragments by Chemical Synthesis

Novel reversible selective inhibitor of nuclear export shows that CRM1 is a target in colorectal cancer cells

Contact Info

Product

Resources

About