Chemokine signaling links cell cycle progression and cilia formation for left-right symmetry breaking

Abstract

23 Zebrafish dorsal forerunner cells (DFCs) undergo vigorous proliferation during 24 epiboly and then exit cell cycle to generate Kupffer's vesicle (KV), a ciliated organ 25 necessary for establishing left-right (L-R) asymmetry. DFC proliferation defects are 26 often accompanied by impaired cilia elongation in KV, but the functional and 27 molecular interaction between cell-cycle progression and cilia formation remains 28 unknown. Here we show that chemokine receptor Cxcr4a is required for L-R laterality 29 by controlling DFC proliferation and KV ciliogenesis. Functional analysis revealed 30 that Cxcr4a accelerates G1/S transition in DFCs and stabilizes Foxj1a, a master 31 regulator of motile cilia, by stimulating Cyclin D1 expression through ERK1/2 32 signaling. Mechanistically, Cyclin D1-CDK4/6 drives G1/S transition during DFC 33 proliferation and phosphorylates Foxj1a, thereby disrupting its association with 34 Psmd4b, a 19S regulatory subunit. This prevents the ubiquitin-independent 35 proteasomal degradation of Foxj1a. Our study uncovers a role for Cxcr4 signaling in 36 L-R patterning and provides fundamental insights into the molecular linkage between 37 cell-cycle progression and ciliogenesis. 38 39 D1-CDK4/6, Foxj1a, Psmd4b, ubiquitin-independent proteasomal degradation.41 42 43 44 3 Author summary 45 During the organogenesis of zebrafish L-R organizer named KV, DFCs 46 proliferate rapidly during epiboly and then exit the cell cycle to differentiate into 47 ciliated epithelial KV cells. Cell cycle defects in DFCs are often accompanied by an 48 alteration in KV cilia elongation. However, whether the cell cycle and cilia formation 49 are mechanistically linked remains as an open question. In this study, we report that 50 Cxcr4 signaling is required for DFC proliferation and KV ciliogenesis. We reveal that 51 Cxcl12b/Cxcr4a signaling activates ERK1/2, which then promotes Cyclin D1 52 expression. Cyclin D1-CDK4/6 accelerates the G1/S transition in DFCs, while also 53 facilitates cilia formation via stabilization of Foxj1a. Notably, Foxj1 undergoes 54 proteasomal degradation via Ub-independent pathway during KV organogenesis. Our 55 study further demonstrates that CDK4 phosphorylates and stabilizes Foxj1a by 56 disrupting its association with Psmd4b, a 19S regulatory subunit. In summary, 57 Cxcl12b/Cxcr4a chemokine signaling links cell cycle progression and cilia formation 58 for L-R symmetry breaking via regulating Cyclin D1 expression. 59 60 61 62 63 64 65 66 67 Vertebrates exhibit striking left-right (L-R) asymmetries in the structure and 68 position of their cardiovascular and gastrointestinal systems. Initially, early embryos 69 develop symmetrically along the prospective body midline. This embryonic symmetry 70 is broken during somite stages when an asymmetric fluid flow is generated by motile 71 cilia within the L-R organizer (LRO), a transient structure located at the posterior end 72 of the notochord [1]. Specifically, in zebrafish, the ciliated LRO is referred to as 73 Kupffer's vesicle (KV), which ...