Dictyostelium discoideum transits from growth phase to developmental phase when nutrients are depleted. D. discoideum under a starved condition aggregates and forms a cell mass called mound within 12 h and then forms a fruiting body. The nutrients critical for the transition are amino acids. Thus, D. discoideum is thought to have a mechanism for sensing amino acid starvation. We examined in the current study the function of Sir2A in the initiation of development. The wild type initiated development and formed mounds within 12 h even in the presence of 1 ~ 3 mg/ml amino acids, while the null mutant did not form mounds within 12 h in the presence of 2 mg/ml amino acid. These results suggest that the null mutant is less sensitive to starvation than is the wild type and that Sir2A plays an important role in the recognition of amino acid starvation.
Linear DNA molecules injected into the macronucleus of Paramecium caudatum became integrated into genomic DNA. The plasmid harboring fusion gene of P. caudatum histone H2B and a yellow fluorescent protein (YFP) named PcVenus was linearized and microinjected into the macronucleus of P. caudatum. Southern blots of total cellular DNA from three fluorescence-positive transformant clones probed with the PcVenus sequence revealed that the injected DNA had become randomly inserted into the chromosome. Free linear monomers or multimers of injected DNA molecules as seen in P. primaurelia and P. tetraurelia were not evident even though the plasmid possessed extant telomeric repeats at its both extremities. We also cloned fragments containing the integration site of genomic DNA from transformant cells by plasmid rescue. Sequence analysis of the flanking DNA confirmed random insertion of the linear plasmid into the chromosomal DNA with extant telomere repeats at the fusion junction. Therefore, P. caudatum maintains introduced DNA in a unique manner by non-homologous or illegitimate, rather than homologous recombination.
The soil-dwelling eukaryotic microorganism Dictyostelium discoideum encodes a protein bearing SNTK, SNN and KTG sequences (DDB0168572) and a protein bearing SISK and HTG sequences (DDB0233562). These sequences are similar to SXXK, S(Y)XN and K(H) T(S)G sequences, which are motifs conserved in penicillin-binding proteins and beta-lactamases. RT-PCR analysis showed that D. discoideum expressed genes for DDB0168572 and DDB0233562 in the growth phase and had decreased expression levels of those genes in the developmental phase. A mutant in which the gene for DDB0168572 was disrupted grew and developed as did the parental strain, indicating that DDB0168572 is dispensable for D. discoideum. Loss of DDB0233562 is thought to be fatal to D. discoideum because no gene-disruption mutant was established.