Synthetic Genetic Controller Circuits to Reprogram Cell Fate
Synthetic Genetic Controller Circuits to Reprogram Cell Fate
批准号:
9367460
负责人:
JAMES J COLLINS
金额:
$63.2万
依托单位国家:
美国
项目类别:
财政年份:
2017
资助国家:
美国
项目状态:
已结题
起止时间:
2017-09-01 至 2021-05-31
关键词:
AddressAntibodiesBCL1 OncogeneBMI1 geneBackBedsBenchmarkingBiologicalBiologyBlood CellsBlood PlateletsBostonCD34 geneCell Differentiation processCellsCommunitiesDataDoseElectrical EngineeringEngineeringEpigenetic ProcessFeedbackFibroblastsFluorescenceFutureGeneticGoalsHematopoietic stem cellsHumanInstitutesKnowledgeLabelLeadMeasuresMechanicsMethodsMicroRNAsModalityPatientsPediatric HospitalsPlatelet TransfusionProcessProtocols documentationRNARegenerative MedicineRegulator GenesResearchScientistStem cell transplantStem cellsStudy modelsSystemTechniquesTestingTranscriptional RegulationTransfectionUmbilical Cord BloodVisionbasec-myc Genescell injurycell typeclinical translationclinically relevantcomputer sciencedesignfeedingfetalimprovedin vivoinduced pluripotent stem cellinsightmathematical analysismathematical modelnovelnovel strategiesoverexpressionpluripotencypredictive modelingpreventprogramspromoterresponsesuccesstooltranscription factortranscriptome sequencing
中文摘要
合成遗传反馈控制器电路重新编程细胞命运
PI:Domitilla Del Vecchio 1;4
共同PI:James J. Collins 2;4;5;6,Thorsten Schlaeger 7和罗恩Weiss 2;3;4
1麻省理工学院机械工程系; 2麻省理工学院生物工程系
3麻省理工学院电子工程与计算机科学系
4麻省理工学院合成生物学中心; 5麻省理工学院和哈佛大学布罗德研究所; 6威斯康星研究所
7干细胞移植项目,波士顿儿童医院
项目摘要
在过去的十年里,干细胞领域有了重大发现,证明了干细胞的命运
与传统观点相反,终末分化细胞可以回复到多能性,
直接转化为其他分化的细胞类型。突然间,再生医学的新方法似乎
触手可及:丢失或受损的细胞可以被患者特异性重编程细胞替代,从而提供
任何所需类型的需求、兼容、高质量电池。为了实现这一愿景,科学界已经做出了
为建立强大而有效的细胞命运重编程方案做出了巨大努力。这些协议
主要基于合适转录因子(TF)的先验固定(prefixed)异位过表达,
这种过度表达可能引发基因调控网络(GRNs)状态之间的转换,
参与细胞命运的决定。然而,尽管十年来取得了显著进展,但这些协议的效率仍然很低,
低,所产生的细胞的质量通常是不令人满意的,并且许多潜在有用的直接细胞命运转化仍然存在。
似乎不可能。这些问题对人类诱导多能干细胞的实际应用构成了巨大的障碍,
细胞(hiPSC)和转分化细胞在再生医学中的应用。
可以说,我们能够准确和精确地将GRNs的TF浓度控制在所需范围内是至关重要的
细胞命运重编程的成功不幸的是,目前基于预固定TF过表达的方案,
没有展现出这种关键的能力。为了解决这个问题,我们提出了一种全新的细胞命运方法,
在这个项目中的重编程:我们用TF的反馈过表达取代了预先固定的过表达,
用体内合成遗传反馈控制器电路实现。在这个回路中,过表达水平不是
先验固定,并根据期望和实际TF浓度之间的差异进行调整。因此,
准确和精确地将TF的浓度控制到所需的值,独立于内源性GRN,
调节这些TF。我们的研究计划首先集中在hiPSC重编程作为一个测试平台,以评估的贝内,
我们的方法和第二个关于hiPSC定向分化为血小板作为直接临床相关应用。
具体而言,在AIM 1中,我们建议系统地研究多能性的prefixed过表达的有效性。
用于hiPSC重编程的TF。在AIM 2中,我们提出了构建和测试合成遗传反馈控制器
同时实现多个TF的反馈过表达的电路。在AIM 3中,我们将利用
用于人hiPSC重编程和hiPSC定向分化的合成遗传反馈控制器电路
变成血小板该项目将大大提高细胞产品的重编程效率,
非常类似于目标细胞类型,在未来,在细胞转换,今天似乎是不可能的。更广泛地说,
我们的合成遗传反馈控制器将使科学家和从业者能够使用一种新的工具来精确地控制
TF的任何内源性GRNs的浓度,特别是参与细胞命运决定的那些GRNs的浓度。
1
英文摘要
Synthetic genetic feedback controller circuits to reprogram cell fate
PI: Domitilla Del Vecchio1;4
co-PIs: James J. Collins2;4;5;6, Thorsten Schlaeger7, and Ron Weiss2;3;4
1Department of Mechanical Engineering, MIT; 2Department of Biological Engineering, MIT
3Department of Electrical Engineering and Computer Science, MIT
4Synthetic Biology Center, MIT; 5Broad Institute of MIT & Harvard; 6The Wyss Institute
7 Stem Cell Transplantation Program, Boston Children's Hospital
PROJECT SUMMARY
The past decade has seen monumental discoveries in the stem cell field, with demonstrations that the fate of a
terminally differentiated cell, contrary to what was traditionally believed, could be reverted back to pluripotency or
directly converted to other differentiated cell types. All of a sudden, new approaches to regenerative medicine seem
within reach: lost or damaged cells could be replaced by patient-specific reprogrammed cells, thus providing on-
demand, compatible, high-quality cells of any required type. To meet this vision, the scientific community has made
tremendous efforts toward establishing robust and efficient protocols for cell fate reprogramming. These protocols are
largely based on a priori fixed (prefixed) ectopic overexpression of suitable transcription factors (TFs), with the rationale
that this overexpression could trigger transitions among the states of the gene regulatory networks (GRNs) that take
part in cell fate determination. Yet, despite a decade of remarkable progress, the efficiency of these protocols remains
low, the quality of produced cells is often unsatisfactory, and many potentially useful direct cell fate conversions still
seem impossible. These issues pose a formidable obstacle to the practical use of both human induced pluripotent stem
cells (hiPSCs) and transdifferentiated cells in regenerative medicine.
Arguably, our ability to accurately and precisely steer the concentrations of GRNs' TFs within desired ranges is critical
to the success of cell fate reprogramming. Unfortunately, current protocols based on prefixed TFs' overexpression have
not demonstrated this critical ability. To address this problem, we propose a completely new approach to cell fate
reprogramming in this project: we replace prefixed overexpression with feedback overexpression of TFs, which we
realize with an in vivo synthetic genetic feedback controller circuit. Within this circuit, the overexpression level is not a
priori fixed and is adjusted based on the discrepancy between desired and actual TF's concentrations. It therefore can
accurately and precisely control TFs' concentrations to desired values, independent of the endogenous GRN that also
regulates these TFs. Our research plan focuses first on hiPSC reprogramming as a test-bed for evaluating the benefit of
our approach and second on directed differentiation of hiPSCs into platelets as a directly clinically relevant application.
Specifically, in AIM 1, we propose to systematically investigate the efficacy of prefixed overexpression of pluripotency
TFs for hiPSC reprogramming. In AIM 2, we propose to construct and test the synthetic genetic feedback controller
circuits that implement feedback overexpression of a number of TFs concurrently. In AIM 3, we will leverage the
synthetic genetic feedback controller circuits for human hiPSC reprogramming and for directed differentiation of hiPSCs
into platelets. This project will result in substantially higher reprogramming efficiencies, in cell products that more
closely resemble the target cell type, and in the future, in cell conversions that today seem not possible. More broadly,
our synthetic genetic feedback controllers will empower scientists and practitioners with a new tool to accurately control
the TFs' concentrations of any endogenous GRNs and, in particular, of those GRNs involved in cell fate determination.
1
期刊论文(0)
专著(0)
科研奖励(0)
会议论文
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