Matrix metalloproteinases in Drosophila wound healing

果蝇伤口愈合中的基质金属蛋白酶

基本信息

批准号：
8893088
负责人：
Andrea W Page-McCaw
金额：
$ 31.17万
依托单位：
VANDERBILT UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-25 至 2017-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8893088
关键词：
Affect Architecture Basement membrane Binding Binding Proteins Biochemical Biochemistry Cancer Patient Cell Culture Techniques Cells Chronic Cicatrix Cleaved cell Complex Data Development Dissection Drosophila genus Epidermis Epithelium Extracellular Matrix Family Genes Genetic Growth Healed Health Human In Vitro Inflammation Knowledge Ligands Link Mammalian Cell Mammals Matrix Metalloproteinase Inhibitor Matrix Metalloproteinases Mediating Methods Microscopy Modeling Molecular Genetics Morphogenesis Mus Organism Pathway interactions Peptide Hydrolases Phenotype Process Property Proteins Proteolysis Proteomics Puncture wound RNA Splicing Receptor Protein-Tyrosine Kinases Role Signal Pathway Signal Transduction Signaling Molecule Site Structure Testing Tissues Transmission Electron Microscopy Ursidae Family Wound Healing base cell motility extracellular genetic analysis healing in vivo innovation loss of function member migration mutant novel receptor repaired research study tool tumor tumor progression wound

项目摘要

DESCRIPTION (provided by applicant): Healing an epidermal wound requires the coordination of tissues and extracellular matrix (ECM), and this coordination is mediated by signaling pathways. Matrix metalloproteinases (MMPs) are proteases that cleave ECM and regulate signaling pathways in vitro, in culture, and during development, but their roles in wound healing are unclear. MMPs are medically important: they are highly upregulated during tumor progression, inflammation, and wound healing. We use a Drosophila model of epidermal wound healing to elucidate MMP function because of its reduced complexity - for example, there are 24 MMPs in mouse, whereas there are only two MMPs in Drosophila, Mmp1 and Mmp2. This organism also offers powerful genetic tools that allow spatio- temporally controlled gain- and loss-of-function phenotypic analyses for nearly every gene. We hypothesize that the Drosophila MMPs form a complex that acts as a global regulator of basement membrane remodeling. This hypothesis is based on genetic analyses of Mmp1, Mmp2, and the MMP binding-protein Timp in wounding and developmental contexts. All are required for healing puncture wounds, for specific aspects of pupal morphogenesis, and for expanding basement membrane during normal growth. Although these genes have other independent phenotypes, the shared phenotypes all point to a fundamental role for the three genes in basement membrane remodeling. As an MMP-Timp trimolecular complex has been described in mammalian cell culture, we hypothesize that Mmp1, Mmp2, and Timp form a similar complex that directs expansion and repair of basement membranes. Supporting this hypothesis, Mmp1's normal localization to the basement membrane and wound margins is lost in Mmp2 and Timp mutants. This hypothesis is novel and exciting: although MMPs are known to cleave ECM, they are considered to degrade ECM, rather than promote its expansion and repair. The MMPs also appear to regulate ERK signaling at the wound margin, with Mmp1 promoting signaling and Mmp2 confining signaling to the margin. These opposing phenotypes suggest another function of the same MMP complex. Aim 1 determines the molecular interactions among Mmp1, Mmp2 and Timp, testing the MMP-complex hypothesis. We will identify binding partners, determine if they co-localize in tissues, and if different Mmp1 splice forms have different partners. Aim 2 determines how MMPs individually and together modify basement membrane, identifying substrates by biochemical methods and an innovative proteomics approach (iTRAQ- TAILS); identifying the tissues controlling remodeling with conditional genetics; and examining the fine structure of mutant basement membrane. Aim 3 determines how the two MMPs oppositely regulate ERK signaling at wounds in vivo, identifying the receptor and ligand, determining the function of ERK signaling in wound healing; and identifying the specific MMP substrate(s) responsible for altering the pathway. To accomplish these aims we have garnered support from a broad team of experts to inform and guide our experiments.

描述（由申请人提供）：治愈表皮伤口需要组织和细胞外基质（ECM）的配位，并且该协调是通过信号通路介导的。基质金属蛋白酶（MMP）是蛋白酶，在体外，培养和发育过程中裂解ECM并调节信号通路，但它们在伤口愈合中的作用尚不清楚。 MMP在医学上很重要：它们在肿瘤进展，炎症和伤口愈合过程中高度上调。由于其复杂性降低，我们使用表皮伤口愈合的果蝇模型来阐明MMP功能 - 例如，小鼠中有24 mMP，而果蝇，MMP1和MMP2中只有两个MMP。该生物还提供了强大的遗传工具，可允许几乎每个基因的时空控制和功能丧失的表型分析。我们假设果蝇MMP形成了一个复杂的复合物，它是基底膜重塑的全球调节剂。该假设基于在伤害和发育环境中MMP1，MMP2和MMP结合蛋白TIMP的遗传分析。愈合穿刺伤口，pupal形态发生的特定方面以及在正常生长过程中扩大基底膜所必需的所有内容。尽管这些基因具有其他独立的表型，但共享表型都表明了这三个基因在基底膜重塑中的基本作用。作为在哺乳动物细胞培养中描述的MMP-TIMP三膜复合物，我们假设MMP1，MMP2和TIMP形成了一种类似的复合物，该复合物指导基底膜的扩张和修复。支持这一假设，MMP1在MMP2和TIMP突变体中损失了MMP1的正常定位和伤口缘。该假设是新颖而令人兴奋的：尽管已知MMP可以切割ECM，但它们被认为会降低ECM，而不是促进其扩张和修复。 MMP似乎还调节了伤口缘处的ERK信号传导，MMP1促进信号传导和MMP2将信号限制在缘。这些相反的表型提出了同一MMP复合物的另一个功能。 AIM 1确定MMP1，MMP2和TIMP之间的分子相互作用，以测试MMP复合假设。我们将确定结合伴侣，确定它们是否在组织中共定位，以及不同的MMP1剪接形式是否具有不同的伴侣。 AIM 2确定MMP如何单独修改基底膜，通过生化方法和创新的蛋白质组学方法（ITRAQ-TAILS）识别底物；鉴定控制有条件遗传学重塑的组织；并检查突变基底膜的精细结构。 AIM 3确定了两个MMP在体内的伤口对ERK信号的反对如何识别受体和配体，从而确定ERK信号在伤口愈合中的功能；并确定负责改变途径的特定MMP底物。为了实现这些目标，我们获得了一群专家团队的支持，以告知和指导我们的实验。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A scar-like lesion is apparent in basement membrane after wound repair in vivo.

DOI：
10.1016/j.matbio.2018.07.004
发表时间：
2018-12
期刊：
Matrix biology : journal of the International Society for Matrix Biology
影响因子：
0
作者：
Ramos-Lewis W;LaFever KS;Page-McCaw A
通讯作者：
Page-McCaw A

A matrix metalloproteinase mediates long-distance attenuation of stem cell proliferation.

DOI：
10.1083/jcb.201403084
发表时间：
2014-09-29
期刊：
The Journal of cell biology
影响因子：
0
作者：
Wang X;Page-McCaw A
通讯作者：
Page-McCaw A

Drosophila Ninjurin A induces nonapoptotic cell death.

DOI：
10.1371/journal.pone.0044567
发表时间：
2012
期刊：
PloS one
影响因子：
3.7
作者：
Broderick S;Wang X;Simms N;Page-McCaw A
通讯作者：
Page-McCaw A

Both Drosophila matrix metalloproteinases have released and membrane-tethered forms but have different substrates.

DOI：
10.1038/srep44560
发表时间：
2017-03-16
期刊：
Scientific reports
影响因子：
4.6
作者：
LaFever KS;Wang X;Page-McCaw P;Bhave G;Page-McCaw A
通讯作者：
Page-McCaw A

Drosophila MMP2 regulates the matrix molecule faulty attraction (Frac) to promote motor axon targeting in Drosophila.

DOI：
10.1523/jneurosci.4811-10.2011
发表时间：
2011-04-06
期刊：
The Journal of neuroscience : the official journal of the Society for Neuroscience
影响因子：
0
作者：
Miller CM;Liu N;Page-McCaw A;Broihier HT
通讯作者：
Broihier HT