EGR1 and EGR2 Involvement in Vertebrate Tendon Differentiation

Abstract

The molecules involved in vertebrate tendon formation during development remain largely unknown. To date, only two DNA-binding proteins have been identified as being involved in vertebrate tendon formation, the basic helix-loop-helix transcription factor Scleraxis and, recently, the Mohawk homeobox gene. We investigated the involvement of the early growth response transcription factors Egr1 and Egr2 in vertebrate tendon formation. We established that Egr1 and Egr2 expression in tendon cells was correlated with the increase of collagen expression during tendon cell differentiation in embryonic limbs. Vertebrate tendon differentiation relies on a muscle-derived FGF (fibroblast growth factor) signal. FGF4 was able to activate the expression of Egr genes and that of the tendon-associated collagens in chick limbs. Egr gene misexpression experiments using the chick model allowed us to establish that either Egr gene has the ability to induce de novo expression of the reference tendon marker scleraxis, the main tendon collagen Col1a1, and other tendon-associated collagens Col3a1, Col5a1, Col12a1, and Col14a1. Mouse mutants for Egr1 or Egr2 displayed reduced amounts of Col1a1 transcripts and a decrease in the number of collagen fibrils in embryonic tendons. Moreover, EGR1 and EGR2 trans-activated the mouse Col1a1 proximal promoter and were recruited to the tendon regulatory regions of this promoter. These results identify EGRs as novel DNA-binding proteins involved in vertebrate tendon differentiation by regulating type I collagen production.Vertebrate tendons are specialized dense connective tissues mainly composed of collagens that connect muscle to bone.Tendon repair following injuries or during aging is a clinical challenge because tendons are only repaired slowly and partially. The establishment of new strategies for tendon repair is prevented by a limited understanding of tendon development and consequently awaits a better understanding of tendon development. An important goal is to understand the molecular mechanisms involved in regulating collagen expression, production, and assembly during tendon development to be able to design new therapies for tendon repair.Tendons consist of elongated fibroblasts named tenocytes that produce an abundant extracellular matrix composed of Ͼ90% collagen. The major collagen component of mature tendon is type I collagen, a heterotrimeric molecule composed of two ␣1 chains and one ␣2 chain that are genetically distinct and encoded by Col1a1 and Col1a2, respectively (1-3). Type I collagen molecules self-assemble into highly organized parallel collagen fibrils and then into fibers, which provide the tensile strength of tendon. Other collagens, such as the fibrillar collagens III and V and the nonfibrillar collagens named FACITs (fibril-associated collagens with interrupted triple helices), and collagens XII and XIV are important for collagen fibril formation, growth, and integrity of tendons. In addition to collagens, tendons contain various matrix components contributing to the prop...

Three-Dimensional Culture and Transforming Growth Factor Beta3 Synergistically Promote Tenogenic Differentiation of Equine Embryo-Derived Stem Cells

“…More robust markers of tenocytes include Scleraxis (SCX), Tenomodulin (TNMD), Tenascin-C (TNC), Cartilage oligomeric matrix protein (COMP), and Thrombospondin-4 (THBS4), which are not only collectively expressed in tendons, but individually also present in other tissue types. [22][23][24][25][26] Developmental studies have also implicated genes such as Six1/2, Eph-A4, Eya1/2, Egr1/2, and Mohawk (Mkx), 25,[27][28][29][30] although it has not been established whether these genes show temporally restricted expression in tendon tissue.…”

Transcriptomic analysis of mouse limb tendon cells during development

“…In addition to the Scx and Tnmd tendon markers, a series of matrix proteins has been described as being expressed or/and associated with tendon development, which we previously attempted to list (Edom-Vovard and Duprez, 2004). The mRNA relative expression of the main tendon collagen, Col1a1, and that of the tendon-associated collagens Col3a1, Col5a1, Col6a1, Col12a1 and Col14a1 was enhanced in tendon cells between E11.5 and E14.5 (Lejard et al, 2011). This is consistent with the high enrichment score of the 'collagen' biological process during tendon development (Table 2).…”

“…SCX has been shown to be required and sufficient for Tnmd expression (Docheva et al, 2005;Shukunami et al, 2006;Murchison et al, 2007). Two other DNA-binding proteins, namely the zinc-finger protein early growth response 1 (EGR1) and the homeodomain protein mohawk (MKX), have been shown to be involved in tendon formation (Ito et al, 2010;Liu et al, 2010;Lejard et al, 2011;Guerquin et al, 2013). However, these two transcription factors, although important for Col1a1 transcription in tendons, are not specific to tendons.…”

Signaux moléculaires et mécaniques intervenant dans la différenciation des cellules tendineuses

“…Les cascades molécu-laires régissant la différenciation du tendon ne sont pas aussi bien décrites que celles des cellules musculaires. Toutefois, l'étude du tendon a été rendue possible grâce à la découverte de certains acteurs moléculaires tels que le facteur de transcription Scx [1][2][3], le plus spécifique et précoce dans le dévelop-pement du tendon, ou encore le facteur de transcription mécanosensitif Egr1 [1,2,4,5]. Au cours des dernières années, de nombreuses études ont montré l'importance des forces mécaniques géné-rées par les cellules ou les tissus au cours du déve-loppement ou des processus de différenciation cellulaire [6].…”