This rapid embryonic cell prolif eration creates more than half of C. elegans somatic cells, with the PXD101 majority of cell divisions being completed in the first half of embryogenesis. Thus, co expres sion of SAC genes in the rapidly dividing early embryo nic cells is consistent with the well established role of these genes in cell division. In addition to the activities of SAC gene promoters in the early embryos, we also observed GFP expression in later embryos for all of the spindle checkpoint promoters that we analyzed. The expression patterns in late embryos show GFP expression in the majority of the cells, although the majority of the promoter constructs tend to confer more localized GFP expression, as exem plified for mdf 2.

Together, the expected promoter activities of SAC genes during embryogenesis, show that the promoters used for our analysis are appropriate. SAC promoters drive tissue specific gene expression later in development Rapid cell proliferation occurs in all four larval stages especially in the second larval stage of develop ment in C. elegans when many somatic cells are nferred by SAC gene promoters was detected at all four larval stages. Unlike embryonic expression, spatio temporal analysis revealed that postembryonic expres sion of SAC genes is generally restricted to specific cells and tissues types. For example, mdf 2 promoter drives GFP expression in seams cells, gut cells, and some additional tissue types at all larval stages. In contrast, mdf 1internal and rod 1 promoters drive GFP expression spe cifically in gut cells after embryogenesis.

Unlike mdf 2, mdf 1 and rod 1 promoters, hcp 1 pro moter was found to be active in the majority, but not all, tissues analyzed, including dorsal ventral nerve cord, head tail body neurons and many other tissue types. Thus, postembryonic spatial analysis revealed distinct, yet overlapping, tissue specific expression of SAC genes during larval development. Unexpectedly, we also observed tissue specific expres sion of SAC genes at late larval and adult stage. Since there are no cell divisions during late L4 and at adulthood except for the divisions in somatic gonads that lead to oocyte development, our observations suggest that SAC genes are expressed in non proliferating cells in C. elegans. Similar to larval expression profiles, tissue specific expression is observed in adult animals as well.

For example, as in larvae, mdf 2 promoter drives GFP expression in seam cells and hypodermis, gut cells, pharynx, and vulva. The expression pat terns detected in adult tissues Brefeldin_A further support the striking co expression of the checkpoint genes in hypodermal seam cells and intestine that we observed in larval stages. Absence of MDF 2 results in aberrant number and alignment of seam cell nuclei We were interested in testing whether absent or non functional SAC would cause aberrant postembryonic seam cell development. For this analysis, we chose mdf 2.