Contribution of cell proliferation to axial elongation in the red flour beetle Tribolium castaneum

Most arthropods generate their posterior bodies by adding segments periodically, as the embryo grows, from a posteriorly located region called the segment addition zone. This mode of segmentation is shared with vertebrates and relies on oscillatory mechanisms, where the temporal periodicity of a clock is translated into repetitive spatial patterns. This ordered anterior-to-posterior pattern is achieved at the same time as the tissue elongates, opening the question of the functional coordination between the mechanisms of segmental patterning and posterior growth. The study of these processes in different arthropods has played an important role in unravelling some of the molecular mechanisms of segment formation. However, the behavior of cells during elongation and how cellular processes affect this segmental patterning has been poorly studied. Cell proliferation together with cell rearrangements are presumed to be the major forces driving axis elongation in the red flour beetle Tribolium castaneum. However, there still no strong evidence about the role and distribution of cell proliferation within the embryo. In this study, we propose to address these questions by using whole embryo cultures and pharmacological manipulation. We show that considerable cell proliferation occurs during germband elongation, measured by incorporation of the nucleoside analog of thymidine 5-Ethynyl-2'-deoxyuridine, EdU. Moreover, proliferating cells appeared to be spread along the elongating embryo with a posterior bias at early segmentation. In addition, when we blocked cell division, treated germbands were always shorter than controls and in some cases not able to fully elongate, even when control embryos already started to retract and leg buds are evident. Finally, we found that the absence of cell proliferation has no apparent effect on segmental patterning, as evidenced by Tc-engrailed (Tc-en) gene expression.