Defining the mechanisms regulating MLLT3 expression in human hematopoietic stem cells
定义人类造血干细胞中 MLLT3 表达的调节机制
基本信息
- 批准号:10113603
- 负责人:
- 金额:$ 28.08万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-03-16 至 2022-12-31
- 项目状态:已结题
- 来源:
- 关键词:BindingBioinformaticsBiologyBlood CellsCRISPR/Cas technologyCell Culture TechniquesChIP-seqChromatinClinicalDataDevelopmentEVI1 geneEngineeringEngraftmentEnhancersEpigenetic ProcessExperimental ModelsFetal LiverFutureGene ExpressionGenerationsGenesGenetic Enhancer ElementGoalsHematological DiseaseHematopoiesisHematopoietic Stem Cell TransplantationHematopoietic stem cellsHigh-Throughput Nucleotide SequencingHistonesHumanImmunologicsIn VitroIndividualKDM1A geneLeadLentivirus VectorLifeLysineMLLT3 geneMaintenanceMalignant - descriptorMediatingModificationMonitorMusNamesPatientsProtein IsoformsProteinsProtocols documentationRegenerative MedicineRegulationRegulator GenesRegulatory ElementReporterRepressionReverse Transcriptase Polymerase Chain ReactionRoleSavingsSecond Pregnancy TrimesterTestingTetanus Helper PeptideTimeTranscriptTranscription Initiation SiteTranscriptional Elongation FactorsUmbilical Cord Bloodaptamerbaseblood formationcell typeepigenome editinggain of functionhematopoietic stem cell expansionhematopoietic stem cell self-renewalhematopoietic tissuehemogenic endotheliumhigh throughput analysishuman RNA sequencinghuman embryonic stem cellimprovedin vivoinsightnovelnovel strategiesoverexpressionp300/CBP-Associated Factorpopulation basedprogramsrecruitself-renewalsingle-cell RNA sequencingstem cell functionstem cell therapystemnesssuccesstransplantation therapy
项目摘要
SUMMARY
Hematopoietic stem cells (HSCs) sustain life-long blood formation due to their ability to self-renew and
differentiate into all mature blood cell types (referred to as “stemness”). Transplantation of HSC-containing grafts
is a life-saving therapy for multiple blood disorders; however, shortage of immunologically matched donors limits
the number of patients that can be treated. Expansion/generation of human HSCs in culture would greatly
improve transplantation therapy but has been unsuccessful due to poor understanding of the underlying biology
of HSC stemness. We identified MLLT3 as HSC regulator that is highly enriched in human HSC at all stages
once they have emerged from AGM and expand in the fetal liver (FL); however, MLLT3 expression declines in
cultured HSC. Loss and gain of function studies on human FL and cord blood (CB) HSCs showed that MLLT3 is
critical for their self-renewal, and when its expression is restored, it enables HSC expansion in culture by
protecting their “stemness” program. Importantly, FL and CB HSC expanded in culture showed 10-30 fold
increase in human engraftment in NSG mice, and ability to sustain the HSPC compartment and multilineage
hematopoiesis without malignant transformation or differentiation block. This finding offers an unprecedented
opportunity to understand how human HSC stemness is controlled, and harness MLLT3 for clinical use. So far,
the regulatory mechanisms that govern MLLT3 expression in HSC are completely unknown. Using a combination
of epigenetic studies and bioinformatic approaches, we identified several candidate enhancers in MLLT3 gene
that may regulate its two isoforms. Our data suggests that these enhancers are epigenetically remodeled during
differentiation and silenced during HSC culture, providing new avenues to understand why cultured HSC lose
MLLT3 expression. To understand how MLLT3 expression is regulated in human HSCs, we propose two
complementary approaches: 1) CRISPR/Cas9-mediated epigenome editing to dissect the role of MLLT3
enhancers, 2) combination of bioinformatic and experimental approaches using lentiviral overexpression to
identify MLLT3 upstream regulators. Success in these approaches could help generate/expand human HSCs in
culture and thereby increase the availability of HSCs for transplantation.
概括
造血干细胞 (HSC) 因其自我更新和自我更新的能力而维持终生血液形成。
分化为所有成熟的血细胞类型(称为“干细胞”)。含有 HSC 的移植物的移植
是治疗多种血液疾病的救命疗法;然而,免疫匹配供体的短缺限制了
可以治疗的患者数量。在培养物中扩增/产生人类 HSC 将极大地促进
改善移植疗法,但由于对潜在生物学了解甚少而未能成功
HSC 干性。我们确定 MLLT3 是 HSC 调节因子,在人类 HSC 的各个阶段都高度富集
一旦它们从 AGM 中出现并在胎儿肝脏 (FL) 中扩展;然而,MLLT3 表达下降
培养的HSC。对人 FL 和脐带血 (CB) HSC 的功能丧失和增强研究表明 MLLT3
对于它们的自我更新至关重要,当其表达恢复时,它可以通过以下方式实现 HSC 在培养物中的扩增:
保护他们的“干性”计划。重要的是,FL 和 CB HSC 在培养物中扩增显示 10-30 倍
NSG 小鼠中人类植入的增加以及维持 HSPC 区室和多谱系的能力
造血无恶变或分化障碍。这一发现提供了前所未有的
有机会了解如何控制人类 HSC 干性,并利用 MLLT3 进行临床应用。迄今为止,
HSC 中控制 MLLT3 表达的调节机制完全未知。使用组合
通过表观遗传学研究和生物信息学方法,我们鉴定了 MLLT3 基因中的几个候选增强子
可能调节其两种亚型。我们的数据表明这些增强子在
HSC 培养过程中的分化和沉默,为理解培养的 HSC 为何丢失提供了新途径
MLLT3 表达。为了了解人类 HSC 中 MLLT3 表达的调节方式,我们提出了两种
补充方法:1) CRISPR/Cas9 介导的表观基因组编辑来剖析 MLLT3 的作用
增强子,2) 结合生物信息学和实验方法,利用慢病毒过表达
识别 MLLT3 上游调节器。这些方法的成功可能有助于在以下领域产生/扩展人类造血干细胞:
培养,从而增加用于移植的 HSC 的可用性。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
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Hanna Katri Annikki Mikkola其他文献
Hanna Katri Annikki Mikkola的其他文献
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{{ truncateString('Hanna Katri Annikki Mikkola', 18)}}的其他基金
MYCT1 as a moderator for signaling between human HSC and their niche
MYCT1 作为人类 HSC 与其生态位之间信号传导的调节剂
- 批准号:
10392239 - 财政年份:2022
- 资助金额:
$ 28.08万 - 项目类别:
MYCT1 as a moderator for signaling between human HSC and their niche
MYCT1 作为人类 HSC 与其生态位之间信号传导的调节剂
- 批准号:
10593103 - 财政年份:2022
- 资助金额:
$ 28.08万 - 项目类别:
Mapping human hematopoietic stem cell development
绘制人类造血干细胞发育图谱
- 批准号:
10435434 - 财政年份:2021
- 资助金额:
$ 28.08万 - 项目类别:
Mapping human hematopoietic stem cell development
绘制人类造血干细胞发育图谱
- 批准号:
10633115 - 财政年份:2021
- 资助金额:
$ 28.08万 - 项目类别:
Mapping human hematopoietic stem cell development
绘制人类造血干细胞发育图谱
- 批准号:
9998658 - 财政年份:2021
- 资助金额:
$ 28.08万 - 项目类别:
Defining the mechanisms regulating MLLT3 expression in human hematopoietic stem cells
定义人类造血干细胞中 MLLT3 表达的调节机制
- 批准号:
9766113 - 财政年份:2019
- 资助金额:
$ 28.08万 - 项目类别:
Defining the mechanisms regulating MLLT3 expression in human hematopoietic stem cells
定义人类造血干细胞中 MLLT3 表达的调节机制
- 批准号:
9894797 - 财政年份:2019
- 资助金额:
$ 28.08万 - 项目类别:
Defining the self-renewal program in human hematopoietic stem cells
定义人类造血干细胞的自我更新程序
- 批准号:
8934081 - 财政年份:2014
- 资助金额:
$ 28.08万 - 项目类别:
Defining the Self-Renewal Program in Human Hematopoietic Stem Cells
定义人类造血干细胞的自我更新程序
- 批准号:
10210386 - 财政年份:2014
- 资助金额:
$ 28.08万 - 项目类别:
Defining the Self-Renewal Program in Human Hematopoietic Stem Cells
定义人类造血干细胞的自我更新程序
- 批准号:
10443733 - 财政年份:2014
- 资助金额:
$ 28.08万 - 项目类别:
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