The HMX homeodomain protein MLS-2 regulates cleavage orientation, cell proliferation and cell fate specification in theC. eleganspostembryonic mesoderm

Abstract

The proper formation of a complex multicellular organism requires the precise coordination of many cellular events, including cell proliferation,cell fate specification and differentiation. The C. eleganspostembryonic mesodermal lineage, the M lineage, allows us to study mechanisms coordinating these events at single cell resolution. We have identified an HMX homeodomain protein MLS-2 in a screen for factors required for M lineage patterning. The MLS-2 protein is present in nuclei of undifferentiated cells in the early M lineage and in a subset of head neurons. In the M lineage,MLS-2 activity appears to be tightly regulated at the fourth round of cell division, coincident with the transition from proliferation to differentiation. A predicted null allele of mls-2, cc615,causes reduced cell proliferation in the M lineage, whereas a semi-dominant,gain-of-function allele, tm252, results in increased cell proliferation. Loss or overexpression of mls-2 also affects cleavage orientation and cell fate specification in the M lineage. We show that the increased cell proliferation in mls-2(tm252) mutants requires CYE-1,a G1 cell cycle regulator. Furthermore, the C. elegans Myod homolog HLH-1 acts downstream of mls-2 to specify M-derived coelomocyte cell fates. Thus MLS-2 functions in a cell type-specific manner to regulate both cell proliferation and cell fate specification.

A Zn-finger/FH2-domain containing protein, FOZI-1, acts redundantly with CeMyoD to specify striated body wall muscle fates in theCaenorhabditis eleganspostembryonic mesoderm

“…LG II: hlh-1(cc561ts), hlh-1(cc450)/mIn1 [dpy-10(e128) mIs14] II (Harfe et al, 1998a) LG III: mab-5(e1239) (Kenyon, 1986), fozi-1(tm0563) (gift from Shohei Mitani, Tokyo Women's Medical University School of Medicine, Japan), ceh-20(n2513) (Liu and Fire, 2000), WM31: dpy-17(e164) vab-7(e1562) LG V: him-5(e1467) (Hodgkin et al, 1979) LG X: mls-2(cc615) (Jiang et al, 2005), hlh-8(nr2061) (Corsi et al, 2000) The CB4856 strain (Hodgkin and Doniach, 1997) was used for snip-SNP mapping of fozi-1.…”

“…In screens for mutants with M lineage defects (Jiang et al, 2005;Foehr et al, 2006), four recessive mutations (cc607, cc608, cc609 and cc610) were isolated that failed to complement one another. These alleles define the fozi-1 locus.…”

“…For immunostaining with HLH-1 antibodies, animals were fixed and stained following the protocol of Harfe and colleagues (Harfe et al, 1998a). The following antibodies were used: affinity purified anti-FOZI-1 (CUMC-GP19; 1:50), goat anti-GFP (Rockland Immunochemicals; 1:5000), rat anti-MLS-2 (Jiang et al, 2005) (1:600) and rabbit anti-HLH-1 (Harfe et al, 1998a) (1:400). All secondary antibodies were from Jackson ImmunoResearch Laboratories and used in a dilution of 1:100 to 1:200.…”

“…The homeodomain protein CEH-20 is required for fozi-1 expression in the M lineage In addition to MAB-5, three other transcription factors crucial for proper M lineage development, CEH-20, MLS-2 and HLH-8, are expressed in the M lineage beginning at the 1-M (or the M mesoblast) stage (Harfe et al, 1998b;Yang et al, 2005;Jiang et al, 2005). We asked whether these factors are required for fozi-1 expression in the M lineage.…”

“…These factors include the Hox factor MAB-5 and its co-factor CEH-20 (Liu and Fire, 2000), the HMX homeodomain protein MLS-2 (Jiang et al, 2005), the MyoD homolog HLH-1 (Harfe et al, 1998a) and the Twist ortholog HLH-8 (Harfe et al, 1998b). MAB-5 and CEH-20 directly activate expression of HLH-8 (Liu and Fire, 2000), while MLS-2 regulates the expression of CeMyoD (HLH-1) in the M lineage (Jiang et al, 2005). Among these transcriptional regulators, HLH-1 is required specifically for BWM and CC fate specification in the M lineage (Harfe et al, 1998a).…”