Nitric oxide interacts with the retinoblastoma pathway to control eye development in Drosophila

Abstract

Animal organ development requires that tissue patterning and differentiation is tightly coordinated with cell multiplication and cell cycle progression. Several variations of the cell cycle program are used by Drosophila cells at different stages during development [1] [2]. In imaginal discs of developing larvae, cell cycle progression is controlled by a modified version of the well-characterized mammalian retinoblastoma (Rb) pathway [3] [4], which integrates signals from multiple effectors ranging from growth factors and receptors to small signaling molecules. Nitric oxide (NO), a multifunctional second messenger [5], can reversibly suppress DNA synthesis and cell division [6] [7]. In developing flies, the antiproliferative action of NO is essential for regulating the balance between cell proliferation and differentiation and, ultimately, the shape and size of adult structures in the fly [8] [9] [10]. The mechanisms of the antiproliferative activity of NO in developing organisms are not known, however. We used transgenic flies expressing the Drosophila nitric oxide synthase gene (dNOS1) and/or genes encoding components of the cell cycle regulatory pathways (the Rb-like protein RBF and the E2F transcription factor complex components dE2F and dDP) combined with NOS inhibitors to address this issue. We found that manipulations of endogenous or transgenic NOS activity during imaginal disc development can enhance or suppress the effects of RBF and E2F on development of the eye. Our data suggest a role for NO in the developing imaginal eye disc via interaction with the Rb pathway.

Regulation of multimers via truncated isoforms: a novel mechanism to control nitric-oxide signaling

“…We show that the action of DNOS4 is reciprocal to RBF and additive or synergistic to dE2F, thus acting to relieve the negative effect of DNOS1 on cell proliferation during eye-disc development. Our results support the hypothesis that the DNOS family of proteins acts to modulate the entry of cells into the S phase of the cell cycle in the eye disc by interacting with the Rb pathway (Kuzin et al 2000; present study). RBF or E2F proteins themselves are potential direct targets for NO action, for example, via S-nitrosylation of cysteine residues by endogenous NO or NO donors (Stamler et al 2001); intriguingly, Rb is one of the several proteins identified in an in vivo screen for S-nitrosylated proteins in mammalian cells (Jaffrey et al 2001).…”

“…NO acts as an antiproliferative factor during eye development; pharmacological inhibition of NOS activity results in an increase in the number of cells in the ommatidia (Kuzin et al 1996(Kuzin et al , 2000. We asked whether DNOS4 can act to suppress the antiproliferative action of NO and affect cell division in the developing eye.…”

“…Because NO interacts with the retinoblastoma (Rb) pathway to control cell division in the developing Drosophila eye (Kuzin et al 2000), we wanted to determine whether DNOS4 can act to affect signaling via the Rb pathway in the eye.…”

“…6A,F; see also Du et al 1996a;Du and Dyson 1999). The RBF phenotype is strongly enhanced by the ectopic overexpression of the dNOS1 transgene (Kuzin et al 2000). To test the effects of DNOS4 on the action of RBF, we crossed GMR-DNOS4-FLAG flies (Fig.…”

“…In mammals, these changes include blood-vessel relaxation, immune response, cell cycle control, and neurotransmission. In Drosophila, NO has been implicated in visual-system development, immunity, behavior, response to hypoxia, osmoregulation, and regulation of cell cycle progression during development (Dow et al 1994;Kuzin et al 1996Kuzin et al , 2000Gibbs and Truman 1998;Wingrove and O'Farrell 1999;Nappi et al 2000;DiGregorio et al 2001;Teodoro and O'Farrell 2003).…”