DOI: 10.5061/dryad.zs7h44j86
title: Cellular dataset for: The anterior Hox gene ceh-13 and elt-1/GATA activate the posterior Hox genes nob-1 and php-3 to specify posterior lineages in the C. elegans embryo

description: Hox transcription factors play a conserved role in specifying positional
 identity during animal development, with posterior Hox genes typically
 repressing the expression of more anterior Hox genes. Here, we dissect the
 regulation of the posterior Hox genes nob-1 and php-3 in the nematode C.
 elegans. We show that nob-1 and php-3 are co-expressed in
 gastrulation-stage embryos in cells that previously expressed the anterior
 Hox gene ceh-13. This expression is controlled by several partially
 redundant transcriptional enhancers. Surprisingly, these enhancers require
 ceh-13 for expression, providing an example of an anterior Hox gene
 positively regulating a posterior Hox gene. Several other regulators
 including elt-1/GATA, ceh-20/ceh-40/Pbx, unc-62/Meis, pop-1/TCF,
 ceh-36/Otx and unc-30/Pitx also act positively through nob-1/php-3
 enhancers. We identified defects in both cell position and cell division
 patterns in ceh-13 and nob-1;php-3 mutants, suggesting that these factors
 regulate lineage identity in addition to positional identity. Taken
 together, our results highlight the complexity and remarkable flexibility
 of Hox gene regulation and function.
description: We acquired confocal images with a Leica TCS SP5, Stellaris or
 Nikon A1RSi resonance scanning confocal microscope (67 z planes at 0.5 μm
 spacing and 1.5 minute time spacing, with laser power increasing by 4-fold
 through the embryo depth to account for attenuation of signal with depth).
 Embryos from self-fertilized hermaphrodites were mounted in egg
 buffer/methyl cellulose with 20μm beads as spacers (Bao and Murray, 2011)
 and imaged at 22°C using a stage temperature controller (Brook Industries,
 Lake Villa, IL). We used StarryNite software to automatically annotate
 nuclei and trace lineages, and AceTree software to identify and fix any
 errors from the automated analysis, and quantified reporter expression in
 each nucleus relative to local background (using the “blot” background
 correction technique) as previously described (Bao et al., 2006; Boyle et
 al., 2006; Murray et al., 2006, 2008; Santella et al., 2014). 
description: For each cell (named according to C. elegans convention (Sulston,
 1983)), for each timepoint collected, there is an expression level (blot
 column) and x, y, z position values. Cells continue until the time at
 which they divide or the dataset (or cell) was no longer curated.
 Additional information about the strains and experimental conditions for
 each embryo are found in the related manuscript. Data
 is presented as comma separated value files (.CSV) which can easily be
 opened in Excel, other spreadsheet program, or with a text editor. Each
 file contains the following columns: cellTime (unique identifier based on
 concatanation of cell name and timepoint), cell (name as based on curated
 assignment according to Sulston 1983), time (order of z-stacks collected
 with 1.5 minute time spacing), none (uncorrected reporter intensity),
 global (corrected reporter intensity with "global" methodology),
 local (corrected reporter intensity with "local" methodology),
 blot (orrected reporter intensity with "blot" methodology, USE
 THIS ONE), cross (corrected reporter intensity with "cross"
 methodology), z (z plane, uncorrected for embryo orientation), x (x
 position in microns, uncorrected for embryo orientation), y (y position in
 microns, uncorrected for embryo orientation), size (cell diameter in
 pixels), gweight (intensity of lineage channel). n.b.
 In file names, ceh-13 refers to JIM118, ceh-13_GFP to JIM356, nob-1 to
 RW10896 and nob-1_GFP to JIM284. The CRISPR GFP reporter strains are
 labeled with their strain numbers. Furthermore, the filenames refer to
 -3.4 kb enhancer as e76 or enh76, the -4.3kb enhancer as e85 or enh85, and
 -8.3 kb enhancers as e92 or enh92. RNAi against a gene of interest is
 indicated by a letter "i" tag following the name of the targeted
 gene, which comes after the reporter name. Additional strain information
 can be found in Supplemental Table 1 of the related manuscript.