Post-transcriptional control and kinetic characterization of proline transport in germinating conidiospores of Aspergillus nidulans.
In the filamentous fungus Aspergillus nidulans, L-proline uptake is mediated by the product of the prnB gene which codes for a member of a family of amino acid transporters found both in pro- and eukaryotes. Regulation of prnB gene expression has previously been studied in great detail at the molecular level. However, no studies have addressed possible post-transcriptional controls or the kinetic characterisation of the PrnB transporter. Here we develop a rapid and efficient method for direct uptake measurements of proline in germinating conidiospores of A. nidulans. We make use of this method and Northern blot analyses in parallel to study the regulation of PrnB expression both at the level of prnB message accumulation and at a post-transcriptional level. These studies show that (i) pathway-specific and wide-domain regulatory systems, previously shown to control prnB gene expression in multicellular mycelia, also operate in unicellular conidia committed to germination; and (ii) PrnB activity is regulated in response to the nitrogen source present in the medium and the level of internally accumulated proline or other amino acids. We also characterise kinetically the PrnB transporter and a secondary proline transport system. Our results open new possibilities for studies using unicellular conidiospores of filamentous fungi and constitute a necessary first step for a subsequent structure-function analysis of the PrnB transporter.
Tazebay
U H
UH
Institut de Génétique et Microbiologie, IGM, Université Paris-Sud, Centre d'Orsay, France.
Sophianopoulou
V
V
Cubero
B
B
Scazzocchio
C
C
Diallinas
G
G
D000596:Amino Acids / Q000378:metabolism; D000641:Ammonia / Q000378:metabolism; D001233:Aspergillus nidulans / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D005246:Feedback; D015966:Gene Expression Regulation, Fungal; D007700:Kinetics; D009584:Nitrogen / Q000378:metabolism; D011392:Proline / Q000235:genetics / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000378:metabolism; D013172:Spores, Fungal / Q000378:metabolism*
D000596:Amino Acids; D011500:Protein Synthesis Inhibitors; D000641:Ammonia; D011392:Proline; D009584:Nitrogen