Ammonium transporter C of Dictyostelium discoideum is required for correct prestalk gene expression and for regulating the choice between slug migration and culmination

Abstract

Ammonium transporter C (AmtC) is one of three transporters in Dictyostelium that have been proposed to regulate entry and exit of ammonia in a cell type dependent manner and to mediate ammonia signaling. Previous work demonstrated that disruption of the amtC gene results in a slugger phenotype in which the cells remain as migrating slugs when they should form fruiting bodies. More detailed studies on the null strain revealed that differentiation of prestalk cell types was delayed and maintenance of prestalk cell gene expression was defective. There was little or no expression of ecmB, a marker for the initiation of culmination. Normal expression of CudA, a nuclear protein required for culmination, was absent in the anterior prestalk zone. The absence of CudA within the tip region was attributable to the lack of nuclear localization of the transcription factor STATa, despite expression of adenylyl cyclase A mRNA in the slug tips. Disruption of the histidine kinase gene dhkC in the amtC null strain restored STATa and CudA expression and the ability to culminate. The results suggest that the lack of nuclear translocation of STATa results from low cAMP due to a misregulated and overactive DhkC phosphorelay in the amtC null strain.

Regulation of Ammonia Homeostasis by the Ammonium Transporter AmtA in Dictyostelium discoideum

“…Previous studies have shown that developmental phenotypes in cells lacking AmtC can be rescued by deleting the AmtA protein (62). These studies suggest that AmtC and AmtA antagonistically regulate developmental processes and that AmtA and AmtC function in either ammonium transport or ammonium sensing (20,37,62). In this study, we show that AmtA regulates intracellular ammonium/ammonia levels during growth and development and is critical for the morphogenesis of multicellular aggregates and normal spore formation.…”

Prolyl 4-hydroxylase-1 mediates O2 signaling during development ofDictyostelium

“…1D, Fig. 5C), a master regulator of culmination (Harwood et al, 1992;Kirsten et al, 2005), which suggests they function in the same pathway. P4H1 is expressed throughout the slug before culmination (Fig.…”

“…Previous studies suggested that overhead light attracts the tip aerially, which promotes culmination indirectly (Bonner et al, 1982), although in the present study of stalled pre-culminants, tips appeared to be elevated from the substratum in either light or dark conditions (not shown). The slug-to-fruit switch is also normally regulated by NH 3 (Kirsten et al, 2005), a catabolic by-product of slug metabolism. NH 3 inhibits culmination in P4H1 -cells at lower concentrations than those required to affect normal cells (Fig.…”

“…Protein kinase A (PKA) has been implicated in regulation of the slug-to-fruit switch at culmination (Kirsten et al, 2005). Activation of PKA by overexpression of the catalytic subunit of PKA under its own promoter (PKAcat) partially rescued culmination in 10% O 2 (Fig.…”

“…The slug-to-fruit switch, thought to be mediated by the anterior tip (Smith and Williams, 1980), is subject to regulation by NH 3 , light, O 2 and other external cues by mechanisms that involve at least ten genes (Schindler and Sussman, 1977;Newell et al, 1969;Sandona et al, 1995;Newell and Ross, 1982). Genetic and biochemical studies suggest that an NH 3 -sensor/transporter, 2-component His kinases (DhkA, DhkC and DhkK), a cAMP phosphodiesterase (RegA), protein kinase A, an E3(SCF FbxA )-Ub ligase, transcription factor-like proteins (STATa, STATc, CudA, MybC), and cytoplasmic red-ox proteins are involved in transducing the signals (Harwood et al, 1992;Mohanty et al, 2001;Kirsten et al, 2005;Singleton et al, 2006;Choi et al, 2006;Thomason et al, 2006). Culmination appears to be exquisitely O 2 -dependent in Dictyostelium, requiring O 2 at a level of greater than 10% (Sandona et al, 1995), whereas growth and early development are relatively normal, down to 2.5% (normoxia=21%).…”