Dataset Information

Hemicentin 2 and Fibulin 1 are required for epidermal-dermal junction formation and fin mesenchymal cell migration during zebrafish development.

ABSTRACT: Hemicentin 1 (Hmcn1) and Hemicentin 2 (Hmcn2) belong to the fibulin family of extracellular matrix (ECM) proteins that play pivotal roles during development and homeostasis of a variety of vertebrate tissues. Recently, we have shown that mutations in zebrafish Hmcn1, also called Fibulin 6, lead to massive fin blistering, similar to the defects caused by the Fraser syndrome gene Fras1. In contrast, the role of Hmcn2 during vertebrate development has thus far been uncharacterized. In zebrafish, hmcn2, like fibulin 1 (fbln1), another member of the fibulin family, is predominantly expressed in fin mesenchymal cells and developing somites, contrasting the strict epithelial expression of hmcn1. While antisense morpholino oligonucleotide (MO)-based knockdown of hmcn2 did not yield any discernable defects, hmcn2/fbln1 double knockdown fish displayed blistering in the trunk, pointing to an essential contribution of these proteins from mesodermal sources for proper epidermal-dermal junction formation. In contrast, and unlike hmcn1 mutants, epidermal-dermal junctions in the fin folds of hmcn2/fbln1 double knockdown fish were only moderately affected. Instead, they displayed impaired migration of fin mesenchymal cells into the fin folds, pointing to a crucial role of Hmcn2 and Fbln1 to remodel the ECM of the fin fold interepidermal space, which is a prerequisite for fibroblast ingrowth. TEM analyses suggest that this ECM remodeling occurs at the level of actinotrichia, the collageneous migration substrate of mesenchymal cells, and at the level of cross fibers, which resemble mammalian microfibers. This work provides first insights into the role of Hmcn2 during vertebrate development, identifying it as an evolutionary conserved protein that acts in functional redundancy with Fbln1C and/or Fbln1D isoforms to regulate tissue adhesion and cell migration, while extending the current knowledge of the functions of vertebrate Fbln1.

SUBMITTER: Feitosa NM

PROVIDER: S-EPMC3423513 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Hemicentin 2 and Fibulin 1 are required for epidermal-dermal junction formation and fin mesenchymal cell migration during zebrafish development.

Feitosa Natália Martins NM Zhang Jinli J Carney Thomas J TJ Metzger Manuel M Korzh Vladimir V Bloch Wilhelm W Hammerschmidt Matthias M

Developmental biology 20120706 2

Hemicentin 1 (Hmcn1) and Hemicentin 2 (Hmcn2) belong to the fibulin family of extracellular matrix (ECM) proteins that play pivotal roles during development and homeostasis of a variety of vertebrate tissues. Recently, we have shown that mutations in zebrafish Hmcn1, also called Fibulin 6, lead to massive fin blistering, similar to the defects caused by the Fraser syndrome gene Fras1. In contrast, the role of Hmcn2 during vertebrate development has thus far been uncharacterized. In zebrafish, hm ...[more]

PMID: 22771579

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Project description:BackgroundVertebrate limb development involves a reciprocal feedback loop between limb mesenchyme and the overlying apical ectodermal ridge (AER). Several gene pathways participate in this feedback loop, including Fgf signaling. In the forelimb lateral plate mesenchyme, Tbx5 activates Fgf10 expression, which in turn initiates and maintains the mesenchyme/AER Fgf signaling loop. Recent findings have revealed that Tbx5 transcriptional activity is regulated by dynamic nucleocytoplasmic shuttling and interaction with Pdlim7, a PDZ-LIM protein family member, along actin filaments. This Tbx5 regulation is critical in heart formation, but the coexpression of both proteins in other developing tissues suggests a broader functional role.ResultsKnock-down of Pdlim7 function leads to decreased pectoral fin cell proliferation resulting in a severely stunted fin phenotype. While early gene induction and patterning in the presumptive fin field appear normal, the pectoral fin precursor cells display compaction and migration defects between 18 and 24 hours post-fertilization (hpf). During fin growth fgf24 is sequentially expressed in the mesenchyme and then in the apical ectodermal ridge (AER). However, in pdlim7 antisense morpholino-treated embryos this switch of expression is prevented and fgf24 remains ectopically active in the mesenchymal cells. Along with the lack of fgf24 in the AER, other critical factors including fgf8 are reduced, suggesting signaling problems to the underlying mesenchyme. As a consequence of perturbed AER function in the absence of Pdlim7, pathway components in the fin mesenchyme are misregulated or absent, indicating a breakdown of the Fgf signaling feedback loop, which is ultimately responsible for the loss of fin outgrowth.ConclusionThis work provides the first evidence for the involvement of Pdlim7 in pectoral fin development. Proper fin outgrowth requires fgf24 downregulation in the fin mesenchyme with subsequent activation in the AER, and Pdlim7 appears to regulate this transition, potentially through Tbx5 regulation. By controlling Tbx5 subcellular localization and transcriptional activity and possibly additional yet unknown means, Pdlim7 is required for proper development of the heart and the fins. These new regulatory mechanisms may have important implications how we interpret Tbx5 function in congenital hand/heart syndromes in humans.

Project description:The evolution of the tetrapod limb involved an expansion and elaboration of the endoskeletal elements, while the fish fin rays were lost. Loss of fin-specific genes, and regulatory changes in key appendicular patterning genes have been identified as mechanisms of limb evolution, however their contributions to cellular organization and tissue differences between fins and limbs remains poorly understood. During early larval fin development, hoxa13a/hoxd13a-expressing fin fold mesenchyme migrate through the median and pectoral fin along actinotrichia fibrils, non-calcified skeletal elements crucial for supporting the fin fold. Fin fold mesenchyme migration defects have previously been proposed as a mechanism of fin dermal bone loss during tetrapod evolution as it has been shown they contribute directly to the fin ray osteoblast population. Using the nitroreductase/metronidazole system, we genetically ablated a subset of hoxa13a/hoxd13a-expressing fin fold mesenchyme to assess its contributions to fin development. Following the ablation of fin fold mesenchyme in larvae, the actinotrichia are unable to remain rigid and the median and pectoral fin folds collapse, resulting in a reduced fin fold size. The remaining cells following ablation are unable to migrate and show decreased actinodin1 mesenchymal reporter activity. Actinodin proteins are crucial structural component of the actinotrichia. Additionally, we show a decrease in hoxa13a, hoxd13a, fgf10a and altered shha, and ptch2 expression during larval fin development. A continuous treatment of metronidazole leads to fin ray defects at 30dpf. Fewer rays are present compared to stage-matched control larvae, and these rays are shorter and less defined. These results suggest the targeted hoxa13a/hoxd13a-expressing mesenchyme contribute to their own successful migration through their contributions to actinotrichia. Furthermore, due to their fate as fin ray osteoblasts, we propose their initial ablation, and subsequent disorganization produces truncated fin dermal bone elements during late larval stages.

Dataset Information

Hemicentin 2 and Fibulin 1 are required for epidermal-dermal junction formation and fin mesenchymal cell migration during zebrafish development.

Publications

Hemicentin 2 and Fibulin 1 are required for epidermal-dermal junction formation and fin mesenchymal cell migration during zebrafish development.

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