Organizer restriction through modulation of Bozozok stability by the E3 ubiquitin ligase Lnx-like.
The organizer anchors the primary embryonic axis, and balance between dorsal (organizer) and ventral domains is fundamental to body patterning. LNX (ligand of Numb protein-X) is a RING finger and four PDZ domain-containing E3 ubiquitin ligase. LNX serves as a binding platform and may have a role in cell fate determination, but its in vivo functions are unknown. Here we show that Lnx-l (Lnx-like) functions as a critical regulator of dorso-ventral axis formation in zebrafish. Depletion of Lnx-l using specific antisense morpholinos (MOs) caused strong embryonic dorsalization. We identified Bozozok (Boz, also known as Dharma or Nieuwkoid) as a binding partner and substrate of Lnx-l. Boz is a homeodomain-containing transcriptional repressor induced by canonical Wnt signalling that is critical for dorsal organizer formation. Lnx-l induced K48-linked polyubiquitylation of Boz, leading to its proteasomal degradation in human 293T cells and in zebrafish embryos. Dorsalization induced by Boz overexpression was suppressed by raising the level of Lnx-l, but Lnx-l failed to counteract dorsalization caused by mutant Boz lacking a critical motif for Lnx-l binding. Furthermore, dorsalization induced by depletion of Lnx-l was alleviated by attenuation of Boz expression. We conclude that Lnx-l modulates Boz activity to prevent the invasion of ventral regions of the embryo by organizer tissue. These studies introduce a ubiquitin ligase, Lnx-l, as a balancing modulator of axial patterning in the zebrafish embryo.
Ro
Hyunju
H
Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-2425, USA.
Dawid
Igor B
IB
D000818:Animals; D019521:Body Patterning / Q000187:drug effects; D002460:Cell Line; D051956:Goosecoid Protein / Q000378:metabolism; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D020897:Organizers, Embryonic / Q000187:drug effects / Q000201:enzymology*; D025821:Polyubiquitin / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D055550:Protein Stability / Q000187:drug effects; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination / Q000187:drug effects; D015027:Zebrafish / Q000196:embryology*; D029961:Zebrafish Proteins / Q000172:deficiency / Q000378:metabolism*
D051956:Goosecoid Protein; D018398:Homeodomain Proteins; C543692:Lnx1 protein, zebrafish; D016376:Oligonucleotides, Antisense; D029961:Zebrafish Proteins; C113874:dharma protein, zebrafish; C497995:gsc protein, zebrafish; D025821:Polyubiquitin; D044767:Ubiquitin-Protein Ligases