李劲松
中科院生物化学与细胞生物学研究所研究员,课题组长,博士生导师
http://www.bio360.net/attachments/character/2016/12/148238944846f5a2b2a3beeef1.jpg

研究学习经历:

编辑
2007 年回国即任中科院上海生命科学研究院生物化学与细胞生物学研究所的研究组长,并入选“百人计划”的李劲松,在建组初期也遇上了从博士后到研究组长的困境。在经历了 4 年焦虑的科研常态后,2011-2012 年他同徐国良合作,连续见刊《Nature》《Science》,并首次建立了来自精子细胞的单倍体胚胎干细胞系,2015 年又建立了来自卵子的单倍体胚胎干细胞系。一系列序贯的成果,极大地推动了单倍体干细胞技术的发展,也正如李劲松提到,沉迷于科研,如同“在海边捡贝壳的小孩,只觉得贝壳越捡越多、越看越美,浑然不觉海水已经漫过了脚踝”。

生物 360 精选了李劲松教授在 2015 年发表的 3 篇文章,翻译摘要以供读者更好的了解其工作成果。由于编辑个人翻译水平有限,如有错误感谢您的指正。

1

Zhong C#, Yin Q#, Xie Z#, Bai M#, Dong R#, Tang W, Xing Y, Zhang H, Yang S, Chen L-L, Bartolomei MS, Ferguson-Smith A, Li D, Yang L*, Wu Y*&Li Jinsong*. CRISPR-Cas9-mediated genetic screening in mice with haploid embryonic stem cells carrying a guide RNA library. Cell Stem Cell2015, 17, 221-232.(注:开源 paper 可下载)

在携带指导 RNA 文库的单倍体胚胎干细胞的小鼠中完成 CRISPR-Cas9 介导的遗传筛选

摘要: 小鼠雄激素单倍体胚胎干细胞 (AG-haESCs) 可以支持半克隆 (SC) 胚胎注射到 MII 卵母细胞中的足月发育,因此在遗传修饰中具有潜在的应用价值。然而,半克隆 (SC) 幼崽的出生率极低,从而限制了这种方法的实际应用。 半克隆 (Semi-Cloned) 胚胎是通过注射体细胞核到未去核的卵母细胞中产生的,在半克隆胚胎中,体细胞被用来作为精子的替代物。然而,由于异常的染色体分离,构建的半克隆胚胎在激活后形成了非整倍体而导致胚胎发育受到严重影响,不能发育到后期。在这里,我们研究发现,AG-haESCs 删除了 DMRs(差异性 DNA 甲基化区域) 中调控两个抑制父本印记基因:H19 和 Gtl2,可以有效支持半克隆(SC)幼崽的出生。 使用 CRISPR-Cas9 在体外对这些 DKO-AG-haESC 的遗传操作可以产生具有高效率且多个修饰的半克隆 (SC) 小鼠。此外,用组成型表达的 sgRNA 文库和 Cas9 转染 DKO-AG-haESC 允许功能性诱变筛选。因此,DKO-AG-haESC 是在单代中在小鼠中引入有机体范围 (organism-wide) 突变的有效工具。

2

Bosley KS, Botchan M, Bredenoord AL, Carroll D, Charo RA, Charpentier E, Cohen R, Corn J, Doudna J, Feng G, Greely HT, Isasi R, Ji W, Kim J, Knoppers B, Lanphier E, Li Jinsong, Lovell-Badge R, Martin GS, Moreno J, Naldini L, Pera M, Perry A, Venter JC, Zhang F & Zhou Q. CRISPR germline engineering-the community speaks. Nature Biotechnology 2015, 33, 5, 478-486.

CRISPR 生殖系修饰工程——the community speaks

摘要: 两种主要的生殖系修饰策略——将 CRISPR-Cas9 转染到受精卵中或将 CRISPR-Cas9 注射到生殖干细胞中(然后产生携带校正基因的配子),生殖干细胞的工程化更有前景。在前一种方法中,不是所有产生的幼崽都携带校正后基因型,并且充分说明脱靶效应需要密切关注;而后一种方法允许在产生合子之前筛选配子的存在和基因修饰的保真度。在生殖干细胞介导的基因治疗可以被人类应用之前,至少有三个极为突出的技术障碍亟待解决。以精源干细胞(SSCs)为例,技术难题第一,如何实现人类干细胞系的有效衍生;第二,是否可以从培养的精源干细胞(SSCs)中获得成熟的精子;第三,是否有可能实现人类精源干细胞(SSCs)的有效遗传修饰?在我看来,在生殖细胞中用 CRISPR-Cas9 纠正人类遗传疾病,其路漫漫其修远兮。

3

Liu P, Dou X, Liu C, Wang L, Xing C, Peng G, Chen J, Yu F, Qiao Y, Song L, Wu Y, Yue C, Li Jinsong, Han J, Tang K & Jing N. Histone deacetylation promotes mouse neural induction by restricting Nodal-dependent mesendoderm fate. Nature Communications 2015, 6: 6830.(注:开源 paper 可下载)

组蛋白脱乙酰化通过限制 Nodal 依赖性的中内胚层细胞促进小鼠的神经诱导

摘要: 细胞命运的决定需要外在信号和内在分子(包括转录因子以及表观遗传调控)之间的合作。然而,表观遗传修饰如何调控神经的分化,在很大程度仍然尚未清楚。在这里我们发现,在外胚层阶段短暂的组蛋白脱乙酰化促进了小鼠胚胎干细胞(mESCs)的神经分化。胚胎干细胞(mESCs)中的组蛋白脱乙酰酶 1(HDAC1)缺失,体外实验—部分拟表型为对组蛋白脱乙酰化的抑制;在体实验—在嵌合小鼠胚胎中显示神经组织中的介入减少。机理研究表明,被组蛋白脱乙酰化抑制的节点 Nodal,是组蛋白脱乙酰酶 1(HDAC1)作用的直接靶标。此外,在 E7.0 小鼠胚胎的前外植体中组蛋白乙酰化的抑制导致 Nodal 激活和神经发育抑制。因此,我们的研究揭示了,通过限制来自体外的小鼠前外胚层神经胶质细胞和来自体内的胚胎干细胞(mESCs)的 Nodal 信号,表观遗传组蛋白去乙酰化确保神经分化的内在机制。

如您对李劲松教授的研究非常关注,请您关注即将在 2017 年 3 月 24、25 日在上海举办的 “基因编辑专题研讨会”。李劲松教授将在会议上就他的研究同各位做深入探讨。

拓展阅读:

李劲松:探索“生命往返跑”奥秘

CRISPR 又发力!中科院李劲松造出“人造精子”

李劲松课题组 Cell Research 发表新成果

马上报名参加 基因编辑专题研讨会

代表性论文:

编辑

1. Zhong C#, Yin Q#, Xie Z#, Bai M#, Dong R#, Tang W, Xing Y, Zhang H, Yang S, Chen L-L, Bartolomei MS, Ferguson-Smith A, Li D, Yang L*, Wu Y*&Li Jinsong*. CRISPR-Cas9-mediated genetic screening in mice with haploid embryonic stem cells carrying a guide RNA library.Cell Stem Cell2015, 17, 221-232.

2. Bosley KS, Botchan M, Bredenoord AL, Carroll D, Charo RA, Charpentier E, Cohen R, Corn J, Doudna J, Feng G, Greely HT, Isasi R, Ji W, Kim J, Knoppers B, Lanphier E, Li Jinsong, Lovell-Badge R, Martin GS, Moreno J, Naldini L, Pera M, Perry A, Venter JC, Zhang F & Zhou Q. CRISPR germline engineering-the community speaks. Nature Biotechnology 2015, 33, 5, 478-486. (Feature)

3. Liu P, Dou X, Liu C, Wang L, Xing C, Peng G, Chen J, Yu F, Qiao Y, Song L, Wu Y, Yue C, Li Jinsong, Han J, Tang K & Jing N. Histone deacetylation promotes mouse neural induction by restricting Nodal-dependent mesendoderm fate. Nature Communications 2015, 6: 6830.

4. Qin Y, Qin J, Zhou C, Li Jinsong& Gao W. Generation of embryonic stem cells from mouse adipose-tissue derived cells via somatic cell nuclear transfer. Cell Cycle 2015, 14: 1282-1290.

5. Zhu W, Yao X, Liang Y, Liang D, Song L, Jing N, Li Jinsong& Wang G*. Mediator Med23 deficiency enhances neural differentiation of murine embryonic stem cells through modulating BMP signaling. Development 2015, 142: 465-476. Epub Jan 6. pii: dev.112946.

6. Wu Y#, Zhou H#, Fan X#, Zhang Y#, Zhang M#, Wang Y, Xie Z, Bai M, Yin Q, Liang D, Tang W, Liao J, Zhou C, Liu W, Zhu P, Guo H, Pan H, Wu C, Shi H, Wu L*, Tang F* &Li Jinsong*. Correction of a genetic disease by CRISPR-Cas9-mediated gene editing in mouse spermatogonial stem cells. Cell Res 2015, 25: 67-79. Epub 2014 Dec 5.

7. LianX, Xu J,Li Jinsong& Chien K. Next-generation models of human cardiogenesis via genome editing. Cold Spring Harb Perspect Med 2014 Sep 18;4(12). pii: a013920. doi: 10.1101/cshperspect.a013920.

8. Guo F#, Li X#, Liang D#, Li T#, Zhu P, Guo H, Wu X, Wen L, Gu T, Hu B, Walsh CP, Li Jinsong*, Tang F* & Xu G*. Active and passive demethylation of male and female pronuclear DNA in the mammalian zygote. Cell Stem Cell 2014, 15: 447-458.[Epub ahead of print].

9. Zhang M, Zhou H, Zheng C, Xiao J, Zuo E, Liu W, Xie D, Shi Y, Wu C, Wang H, Li D & Li Jinsong. The roles of testicular c-kit positive cells in de novo morphogenesis of testis. Sci Rep 2014, 4: 5936.

10. Zhu Q, Song L, Peng G, Sun N, Chen J, Zhang T, Sheng N, Tang W, Qian C, Qiao Y, Tang K, Han JD, Li Jinsong& Jing N. The transcription factor Pou3f1 promotes neural fate commitment via activation of neural lineage genes and inhibition of external signaling pathways.Elife 2014, Jun 14:e02224. doi: 10.7554/eLife.02224. [Epub ahead of print].

11. Zhao B, Yang D, Jiang J, Li Jinsong, Fan S, Huang M, Fan Y, Jin Y & Jin Y. Genome-wide mapping of miRNAs expressed in embryonic stem cells and pluripotent stem cells generated by different reprogramming strategies. BMC Genomic 2014, 15: 488.

12. Yu Y, Liang D, Tian Q, Chen X, Jiang B, Chou BK, Hu P Cheng L, Gao P, Li Jinsong& Wang G. Stimulation of somatic cell reprogramming by Eras-Akt-Foxo1 signaling axis. Stem Cells 2014,32(2): 349-363. doi: 10.1002/stem.1447. [Epub ahead of print].

13. Qin Y, Zhou P, Zhou C, Li Jinsong& Gao W. The adipose-derived lineage-negative cells are enriched mesenchymal stem cells and promote limb ischemia recovery in mice. Stem Cells Dev 2014, 23: 363-371.

14. Wu Y#, Liang D#, Wang Y, Bai M, Tang W, Bao S, Yan Z, Li D &Li Jinsong*. Correction of a genetic disease in mouse via use of CRISPR-Cas9. Cell Stem Cell 2013, 13: 659-662.

15. Yang H#, Liu Z#, Ma Y#, Zhong C, Yin Q, Zhou C, Shi L, Cai Y, Zhao H, Wang H, Tang F, Wang Y, Zhang C, Liu X, Lai D, Jin Y*, Sun Q* &Li Jinsong*. Generation of haploid embryonic stem cells from Macaca fascicularis monkey parthenotes. Cell Res 2013, 23: 1187-1200.

16. Qin Y#, Lin J#, Zhou C#, Yin Q, Xie Z, Zhang X, Liu X, Gao W &Li Jinsong*. Mice cloned from white adipose tissue-derived cells. J Mol Cell Biol 2013, 5: 348-350.

17. Jiang J#, Lv W#, Ye X, Wang L, Zhang M, Yang H, Okuka M, Zhou C, Zhang X, Liu L* & Li Jinsong*. Zscan4 promotes genomic stability during reprogramming and dramatically improves the quality of iPS cells as demonstrated by tetraploid complementation. Cell Res2013, 23: 92-106.

18. Zhao S#, Gou LT#, Zhang M#, Zu LD, Hua MM, Hua Y, Shi HJ, Li Y, Li Jinsong, Li DS, Wang ED* & Liu M. piRNA-triggered MIWI ubiquitination and removal by APC/C in late spermatogenesis. Dev Cell 2013, 24: 13-25.

19. Yang H#, Shi L#, Wang B#, Liang D, Zhong C, Liu W, Nie Y, Liu J, Zhao J, Gao X, Li D, Xu G-L*&Li Jinsong*. Generation of genetically modified mice by oocyte injection of androgenetic haploid embryonic stem cells. Cell 2012, 149(3): 605-617.

20. Shi L, Yang H &Li Jinsong*. Haploid embryonic stem cells: an idea tool for mammalian genetic analyses. Protein & Cell 2012, 3: 806-810. (Invited review)

21. Ma Y, Gu J, Li C, Wei X, Tang F, Shi G, Jiang J, Kuang Y, Li Jinsong, Wang Z, Xie X & Jin Y. Human foreskin fibroblast produces interleukin-6 to support derivation and self-renewal of mouse embryonic stem cells.Stem Cell Res Ther 2012, 3: 29.

22. Gu TP#, Guo F#, Yang H#, Wu HP, Xu GF, Liu W, Xie ZG, Shi L, He X, Jin SG, Iqbal K, Shi YG, Deng Z, Szabo PE, Pfeifer GP, Li Jinsong* & Xu GL*. The role of Tet3 DNA dioxygenase in epigenetic reprogramming by oocytes. Nature 2011, 477, 606-610.

23. Jiang J#, Ding G#, Lin J#, Zhang M, Shi L, Lv W, Yang H, Xiao H, Pei G, Li Y, Wu J* &Li Jinsong*. Different developmental potential of pluripotent stem cells generated by different reprogramming strategies. J Mol Cell Biol2011, 3: 197-199.

24. Lin J, Shi L, Zhang M, Yang H, Qin Y, Zhang J, Gong D, Zhang X, Li D &Li Jinsong*. Defects in trophoblast cell lineage account for the impaired in vivo development of cloned embryos generated by somatic nuclear transfer. Cell Stem Cell2011, 8: 371-375.

25. Yang H#, Shi L#, Chen D &Li Jinsong*. Mice generated after round spermatid injection into haploid two-cell blastomeres. Cell Res2011, 21: 854-858.

26. Li X, Zhu L, Yang A, Lin J, Tang F, Jin S, Wei Z, Li Jinsong&Jin Y. Calcineurin-NFAT signaling critically regulates early lineage specification in mouse embryonic stem cells and embryos. Cell Stem Cell 2011 8: 46-58.

27. Wang Y, Chen J, Hu J, Wei X, Qin D, Gao J, Zhang L, Jiang J, Li Jinsong, Liu J, Lai K, Kuang X, Zhang J, Pei D & Xu G. Reprogramming of mouse and human somatic cells by high-performance engineered factors.EMBO Rep 2011, 12: 373-378.

28. Yang H, Shi L, Zhang S, Ling J, Jiang J&Li Jinsong*. High-efficiency somatic reprogramming induced by intact MII oocytes.Cell Res 2010, 20, 1034-42.

29. Li L, Sun L, Gao F, Jiang J, Li C, Gu J, Wei Z, Yang A, Lu R, Ma Y, Tang F, Kwon SW, Zhao Y, Li Jinsong& Jin Y. Stk40 links the pluripotency factor Oct4 to the Erk/MAPK pathway and controls extraembryonic endoderm differentiation. Proc Natl Acad Sci USA 2010, 107(4):1402-1407.

30. Chen T, Yuan D, Wei B, Jiang J, Kang J, Ling K, Gu Y, Li Jinsong, Xiao L& Pei G. E-cadherin-mediated cell-cell contact is critical for induced pluripotent stem cell generation. Stem Cells2010, 28: 1315-1325.

31. Zhu Z, Wang Y, Li X, Wang Y, Xu L, Wang X, Sun T, Dong X, Chen L, Mao H, Yu Y, Li Jinsong, Chen PA&Chen CD. PHF8 is a histone H3K9me2 demethylase regulating rRNA synthesis.Cell Res2010, 20: 794-801.

32. Li C, Yu H, Ma Y, Shi G, Jiang J, Gu J, Yang Y, Jin S, Wei Z, Jiang H, Li Jinsong& Jin Y. Germliine-competent mouse-induced pluripotent stem cell lines generated on human fibroblasts without exogenous leukemia inhibitory factor. PLos One2009, 4: e6724.

33. Huang J, Chen T, Liu X, Jiang J, Jinsong Li, Li D, Liu X, Li W, Kang J& Pei G. More synergetic cooperation of Yamanaka factors in induced pluripotent stem cells than in embryonic stem cells. Cell Res2009, 19: 1127-1138.

34. LiJinsong& Mombaerts P. Nuclear transfer-mediated rescue of the nuclear genome of nonviable mouse cells frozen without cryoprotectant. Biol Reprod 2008 79(4):588-593.

35. Li Jinsong#, Guasch G#, Greco V#, Fuchs E* & Mombaerts P*. Cloned mice from Skin Stem Cells.Proc Natl Acad Sci USA 2007, 104(8): 2738-2743.

36. LiJinsong,Ishii T, Wen D& Mombaerts P. Non-equivalence of cloned and clonal mice. Current Biology2005, 15(18): R756-R757.

37. LiJinsong, IshiiT, FeinsteinP&Mombaerts P. Odorant receptor gene choice is reset by nuclear transfer from mouse olfactory sensory neurons. Nature 2004, 428: 393-398.

38. Han Z, Chen D, Li Jinsong, Sun Q, Wan Q, Kou Z, Rao G, Lei L, Liu Z & Fang S. Mitochondrial DNA heteroplasmy in calves cloned by using adult somatic cell. Mol Reprod Dev2004, 67(2): 207-214

39. Chen D#, Li Jinsong#, Han Z, Lei L, Liu Z, Kou Z, Ma S, Du Q & Sun Q. Somatic cell bovine cloning: efficiency of donor cells from femaleand male using different recipient cows. Chinese Science Bulletin 2003, 48(6): 549-554.

40. Zhu Z, Chen D, Li Jinsong, Lian L, Lei L, Han Z & Sun Q. Rotation of meiotic spindle is controlled by microfilaments in mouse oocytes.Biology of Reproduction 2003, 68: 943-946.

41. Han Z, Chen D, Li Jinsong, Sun Q, Wang P, Du J & Zhang H. Flow cytometric cell-cycle analysis of cultured fibroblasts from the giant panda, Ailuropoda melanoleuca L. Cell Biol Int 2003, 27(4): 349-353

42. Wen D, Yang C, Cheng Y, Li Jinsong, Liu Z, Sun Q, Zhang J, Lei L, Wu Y, Kou Z &Chen D. Comparison of developmental capacity for intra- and interspecies cloned cat (Felis catus) embryos. Mol Reprod Dev 2003, 66(1): 38-45

43. Chen Y, He Z, Liu A, Wang K, Mao W, Chu J, Lu Y, Fang Z, Shi Y, Yang Q, Chen D, Wang M, Li Jinsong, Huang S, Kong X, Shi Y, Wang Z, Xia J, Long Z, Xue Z, Ding W & Sheng H. Embryonic stem cells generated by nuclear transfer of human somatic nuclei into rabbit oocytes.Cell Res2003, 13(4): 251-263.

44. Li Jinsong, Chen D, Han Z, Zhu Z, Wen D, Liu Z, Wang M, Lian L, Du J, Wang P& Zhang H. Serial Nuclear Transfer Improves the Development of Interspecies Reconstructed Giant Panda Embryos. Chinese Science Bulletin2002, 47(6):467-469

45. Chen D, Wen D, Zhang Y, Sun Q, Han Z, Liu Z, Shi P, Li Jinsong, Xianyu J, Lian L, Kou Z, Wu Y, Chen Y, Wang P& Zhang H. Interspeices implantation and mitochondria fate of panda-rabbit cloned embryos. Biol Reprod 2002,67:637-642.

46. Han Z, Chen D, Li Jinsong, Sun Q, Wang P, Huang Y & Du J. The culture of fibroblasts from diaphragm of Giant Panda. In Vitro Cell Dev Biol 2001, 37:644-645

47. Li G, Chen D, Lian L, Sun Q, Wang M, Liu J, Li Jinsong& Han Z. Viable rabbits derived from reconstructed oocytes by germinal vesicle transfer after intracytoplamic sperm injection (ICSI). Mol Reprod Dev2001, 58:180-185

48. Ding B, Shi P, Xianyu J, Zhang Y, Chen D, Sun Q, Li G, Wang M, Liu J, Han Z, Song X, Li Jinsong& Chen Y. Microsatellite DNA analysis prove nucleus of interspecies reconstructed blastocyst coming from that of donor giant panda. Chinese Science Bulletin 2000, 45(20):1883-1885

49. Liu J, Wang M, Lian L, Li jinsong, Gao S, Han Z, Sun Q, Song X, Zhang D, Li Y, Xu Z &Chen D. Nuclear Transfer Using Nonquiescent Adult Fibroblasts from a Bovine Ear. Chinese Science Bulletin1999, 44(21): 1971-1974

50. Chen D, Sun Q, Liu J, Li G, Lian L, Wang M, Han Z, Song X, Li Jinsong, Sun Q, Chen Y, Zhang Y & Ding B. The Giant Panda Somatic Nucleus can dedifferentiate in Rabbit Ooplasm and Support Early Development of the Reconstructed Egg. Science in China 1999, 42(4): 346-353

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