The rare actinomycete Actinoplanes missouriensis forms terminal sporangia containing a few hundred flagellated spores, which can swim in aquatic environments after release from sporangium. However, gene regulation for its characteristic morphological development is largely unknown. Here, we report the functional analysis of an orphan response regulator, TcrA, which is encoded next to the chemotaxis-flagellar gene cluster. The tcrA null (ΔtcrA) mutant formed sporangium, in which sporulation proceeded. However, many distorted spores were produced and some spores ectopically germinated in the mutant sporangia. In addition, spores were hardly released from the mutant sporangia. A comparative RNA-Seq analysis between the wild-type and ΔtcrA strains showed that TcrA upregulated the transcription of more than 263 genes, which were integrated into 185 transcriptional units. In silico searches identified a 21-bp direct repeat sequence, 5'-nnGCA(A/C)CCG-n -GCA(A/C)CCGn-3', as the TcrA box, which was confirmed by electrophoretic mobility shift assays. Finally, we identified 34 transcriptional units as the TcrA regulon. TcrA seems to regulate a few hundred genes through the transcriptional activation of three FliA-family sigma factor genes besides its own regulon. We concluded that TcrA is a global transcriptional activator that controls many aspects of sporangium formation, including flagellar biogenesis, spore dormancy and sporangium dehiscence.

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The rare actinomycete Actinoplanes missouriensis forms sporangia, including hundreds of flagellated spores that start swimming as zoospores after their release. Under conditions suitable for vegetative growth, zoospores stop swimming and germinate. A comparative proteome analysis between zoospores and germinating cells identified 15 proteins that were produced in larger amounts in germinating cells. They include an orthologue of BldD (herein named AmBldD [BldD of A. missouriensis]), which is a transcriptional regulator involved in morphological development and secondary metabolism in Streptomyces. AmBldD was detected in mycelia during vegetative growth but was barely detected in mycelia during the sporangiumforming phase, in spite of the constant transcription of AmbldD throughout growth. An AmbldD mutant started to form sporangia much earlier than the wild-type strain, and the resulting sporangia were morphologically abnormal. Recombinant AmBldD bound a palindromic sequence, the AmBldD box, located upstream from AmbldD. 3=,5=-Cyclic di-GMP significantly enhanced the in vitro DNA-binding ability of AmBldD. A chromatin immunoprecipitation-sequencing analysis and an in silico search for AmBldD boxes revealed that AmBldD bound 346 genomic loci that contained the 19-bp inverted repeat 5=-NN(G/A)TNACN(C/G)N(G/C)NGTNA(C/T)NN-3= as the consensus AmBldDbinding sequence. The transcriptional analysis of 27 selected AmBldD target gene candidates indicated that AmBldD should repress 12 of the 27 genes, including bldM, ssgB, whiD, ddbA, and wblA orthologues. These genes are involved in morphological development in Streptomyces coelicolor A3(2). Thus, AmBldD is a global transcriptional regulator that seems to repress the transcription of tens of genes during vegetative growth, some of which are likely to be required for sporangium formation.IMPORTANCE The rare actinomycete Actinoplanes missouriensis undergoes complex morphological differentiation, including sporangium formation. However, almost no molecular biological studies have been conducted on this bacterium. BldD is a key global regulator involved in the morphological development of streptomycetes. BldD orthologues are highly conserved among sporulating actinomycetes, but no BldD orthologues, except one in Saccharopolyspora erythraea, have been studied outside the streptomycetes. Here, it was revealed that the BldD orthologue AmBldD is essential for normal developmental processes in A. missouriensis. The AmBldD regulon seems to be different from the BldD regulon in Streptomyces coelicolor A3(2), but they share four genes that are involved in morphological differentiation in S. coelicolor A3(2).KEYWORDS gene regulation, rare actinomycete, regulon, sporangium formation, transcriptional factor A ctinomycetes are Gram-positive, mainly soil-inhabiting bacteria. Many genera of actinomycetes show filamentous growth and produce spores, and therefore, actinomycetes are generally characterized by a complex morphological development. For

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The Antarctic minke whale (Balaenoptera bonaerensis) is the most abundant baleen whale species in the Southern Ocean. Quantitative information on prey consumption of whales is useful to understand their feeding ecology and role in the ecosystem. The purposes of this study are 1) to investigate the feeding habit of Antarctic minke whales based on information on prey species, freshness and diurnal change in stomach contents, and 2) to estimate the amount of prey consumed by whales. Estimates are made for whales of different sexual maturity classes as it is expected that the energy requirements vary among them. The analysis is based on the data from whales taken by JARPA (Japanese Whale Research Program under Special Permit in the Antarctic) in a longitudinal sector between 35º E and 145º W, and south of 60º S. Sampling was conducted in the austral summer seasons from 1987/1988 to 2004/2005, mainly in the months from December to March. Daily prey consumption by the whales in each sexual maturity class was estimated using energy-requirement and energy deposition. The whales feed mainly before 05:00 h, which suggest that they cease to feed early in the day. Daily prey consumptions were estimated to be 83.7-325.5 kg, equivalent to 2.7-4.0% of body weight. The mean prey consumptions per capita during feeding season were 7.5 and 16.4 t for immature and mature male, 12.5 and 39.1 t for immature and mature female, respectively. In Area IV (70°-130°E), total prey consumptions of krill by Antarctic minke whales in 1999/2000 and 2001/2002 seasons were estimated to be 0.9 and 1.1 million t, respectively. In Area V (130° E-170° W including the Ross Sea), these estimates in 2000/2001 and 2002/2003 seasons were 3.9 and 4.1 million t, respectively. The estimations of feeding impact on krill resources by Antarctic minke whales in Areas IV and V were from 2.7 to 3.2%, and from 18.2 to 18.9% of krill biomasses, respectively. These results on prey consumption are important input data for the development of ecosystem modeling in the Southern Ocean.

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