Developmental spatiotemporal switching of the germline-specific subunits of heterodimeric NAC protein in Drosophila melanogaster.

The ribosome-associated αβ heterodimeric ubiquitously expressed NAC protein is involved in protein homeostasis in eukaryotes. We previously reported that the germline-specific α and β subunits (gNACαβ) differ from ubiquitously expressed paralogs by the presence of extended intrinsically disordered regions. The embryonic precursors of the germline (pole cells) express both α and β gNAC subunits. CRISPR/Cas9-mediated knockout of the gNAC α- subunit gene resulted in the death of pole cells progenitors, implicating gNAC as an essential component of the germ plasm responsible for the germline development. Immunofluorescence detection was used to track changes in the expression of α- and β-subunits of gNAC during development. The bright fluorescence of the α-subunit in the germarium strongly decreases during oocyte specification and is replaced by the onset of increased β-subunit fluorescence, especially in the posterior part of mature oocytes, in the germ plasm region, where NACα-subunit fluorescent signal is absent. The visualized switches of gNAC subunits presence, as well as detection of separate cytoplasmic fluorescent puncta for α and β gNAC subunits in a germline cells (elongating spermatids in testis or embryonic pole cells), are explained by transient formation the hybrid NAC heterodimers composed of α or β germline subunit and a ubiquitous partner. The formation of hybrids, demonstrated by mass spectrometry analysis in testes, indicates a programed spatiotemporal functioning of both the ubiquitous and germline-specific NAC paralogs to support the germline-specific proteostasis.