A Lineage Analysis of Cells in the Vertebrate Limb

脊椎动物肢体细胞的谱系分析

• 批准号: 6690508
• 负责人: Jennifer H Mansfield
• 金额: $ 3.97万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6690508
• 关键词: biological signal transduction; cell differentiation; chick embryo; connective tissue development; developmental genetics; genetically modified animals; histogenesis; laboratory mouse; lac operon; limbs; mesoderm; polymerase chain reaction; postdoctoral investigator

DESCRIPTION (provided by applicant): Although there has been enormous progress toward understanding the intercellular signals that organize the developing vertebrate limb, practically nothing is known about the lineage relationships between cells that give rise to the various limb tissues, or of the differential fates of cells which produce the organizing signals. Defining such lineage relationships is essential to understanding limb morphogenesis. Here, a twofold strategy is proposed. First, individual cells derived from the limb lateral plate mesoderm (LPM) will be traced, by labeling with a complex library of oligonucleotide-tagged retroviruses, to ask when individual cells and their descendents become committed to particular cell fates and spatial locations. The LPM gives rise to the limb skeleton, connective tissue, smooth muscle and dermis, but how and when these populations diverge is unknown. Second, the progeny of cells in the Zone of Polarizing Activity (ZPA) will be traced. ZPA cells express Sonic Hedgehog (Shh), a secreted signal that polarizes the limb along its A/P axis. Permanent, heritable LacZ expression will be generated in all descendents of the Shh-expressing ZPA using the Cre-recombinase system. Some cells in the posterior limb bud, a region that includes the ZPA, undergo apoptosis while others contribute to digit structures. Whether descendents of the ZPA can reprogram gene expression patterns and contribute to limb structures is unknown. If true, the identification of such dynamic shifts in Shh expression would have profound implications for the mechanism of limb patterning.

描述（由申请人提供）： 尽管在理解组织发育中的脊椎动物肢体的细胞间信号方面已经取得了巨大进展，但实际上对产生各种肢体组织的细胞之间的谱系关系或产生组织信号的细胞的差异命运一无所知。定义这种谱系关系对于理解肢体形态发生至关重要。在这里，提出了双重策略。首先，通过用寡核苷酸标记的逆转录病毒的复杂文库进行标记，将追踪源自肢体侧板中胚层（LPM）的单个细胞，以询问单个细胞及其后代何时决定特定的细胞命运和空间位置。 LPM 产生肢体骨骼、结缔组织、平滑肌和真皮，但这些群体如何以及何时分化尚不清楚。其次，将追踪极化活动区（ZPA）中的细胞后代。 ZPA 细胞表达 Sonic Hedgehog (Shh)，这是一种沿着 A/P 轴极化肢体的分泌信号。使用 Cre 重组酶系统，将在表达 Shh 的 ZPA 的所有后代中产生永久、可遗传的 LacZ 表达。后肢芽（包括 ZPA 的区域）中的一些细胞会发生凋亡，而其他细胞则会形成数字结构。 ZPA 的后代是否可以重新编程基因表达模式并对肢体结构做出贡献尚不清楚。如果属实，那么识别 Shh 表达的这种动态变化将对肢体模式的机制产生深远的影响。