CLICK ON weeks 0 - 40 and follow along every 2 weeks of fetal development
Developmental biology - Genes|
The gene code of growing limbs
The big question in developmental biology for a long time has been: What sets the pace and order of developmental events?
Scientists in the labs of Denis Duboule at EPFL, amd Pierre Fabre at the University of Geneva, have now identified the gene cascade forming the paw and digit 'architecture' in mice. They are Hoxd genes that are developmentally active in various combinations inside "progenitor cells" - cells a step more specialized than stem cells.
Ushing a cutting-edge single-cell RNA sequencing technique, they studied expression patterns in thousands of genes inside individual progenitor cells. Their analysis showed that each cell contains 343 genes, in specific cellular states within the cell nucleus. Many of these states are associated with DNA packaging and affect how a paw's fingers will develop.
Scientists identified a very restricted set of six principal combinations of five Hoxd genes (Hoxd9, 10, 11, 12 and 13) in digit development of mice. Each combination includes one, two, or four genes - the single gene beginning development and more complex (four genes) occurring in later stages.
The pattern by which Hox genes are expressed when "building" the entire body in both mice and humans, has been known and studied for over thirty years. Most studies look at the tissues of developing organs. In contrast, this study is the first to look at Hox in single cells, at higher resolution to see how genes orchestrate the rhythm of development.
"The study shows how architect genes act in concert. They follow a gradual progression in every developing cell in order to generate fully mature arms and hands at the right time and place.
Global analyses of gene expression during development reveal specific transcription patterns associated with the emergence of various cell types, tissues, and organs. These heterogeneous patterns are instrumental to ensure the proper formation of the different parts of our body, as shown by the phenotypic effects generated by functional genetic approaches. However, variations at the cellular level can be observed within each structure or organ. In the developing mammalian limbs, expression of Hox genes from the HoxD cluster is differentially controlled in space and time, in cells that will pattern the digits and the forearms. While the Hoxd genes broadly share a common regulatory landscape and large-scale analyses have suggested a homogenous Hox gene transcriptional program, it has not previously been clear whether Hoxd genes are expressed together at the same levels in the same cells.
We report a high degree of heterogeneity in the expression of the Hoxd11 and Hoxd13 genes. We analyzed single-limb bud cell transcriptomes and show that Hox genes are expressed in specific combinations that appear to match particular cell types. In cells giving rise to digits, we find that the expression of the five relevant Hoxd genes (Hoxd9 to Hoxd13) is unbalanced, despite their control by known global enhancers. We also report that specific combinatorial expression follows a pseudo-time sequence, which is established based on the transcriptional diversity of limb progenitors.
Our observations reveal the existence of distinct combinations of Hoxd genes at the single-cell level during limb development. In addition, we document that the increasing combinatorial expression of Hoxd genes in this developing structure is associated with specific transcriptional signatures and that these signatures illustrate a temporal progression in the differentiation of these cells.
P. J. Fabre, M. Leleu, B. Mascrez, Q. Lo Giudice, J. Cobb and D. Duboule.
EPFL; University of Geneva; Swiss National Science Foundation (Ambizione); and the European Research Council (SystemHox and RegulHox).
This study is a collaboration between the faculties of Science and Medicine of the University of Geneva with EPFL's ISREC Foundation and BBCF.
Return to top of page
Illustration of the Hox gene code stretched over a hand sculpture by Mario Irrarazabal. Each dot is one cell. Colors represent the differentiation order of single-cells. Credit: Sculpture 'and del disierto' by Mario Irrarazabal in the Atacama desert of Chile. Graphic of pseudotime by P. Fabre and
Q. Lo Giudice, University of Geneva.