Regulation of the KNOX-GA Gene Module Induces Heterophyllic Alteration in North American Lake Cress

Plants show leaf form alteration in response to changes in the surrounding environment, and this phenomenon is called heterophylly. Although heterophylly is seen across plant species, the regulatory mechanisms involved are largely unknown. Here, we investigated the mechanism underlying heterophylly in Rorippa aquatica (Brassicaceae), also known as North American lake cress. R. aquatica develops pinnately dissected leaves in submerged conditions, whereas it forms simple leaves with serrated margins in terrestrial conditions. We found that the expression levels of KNOTTED1-LIKE HOMEOBOX (KNOX1) orthologs changed in response to changes in the surrounding environment (e.g., change of ambient temperature; below or above water) and that the accumulation of gibberellin (GA), which is thought to be regulated by KNOX1 genes, also changed in the leaf primordia. We further demonstrated that exogenous GA affects the complexity of leaf form in this species. Moreover, RNA-seq revealed a relationship between light intensity and leaf form. These results suggest that regulation of GA level via KNOX1 genes is involved in regulating heterophylly in R. aquatica. The mechanism responsible for morphological diversification of leaf form among species may also govern the variation of leaf form within a species in response to environmental changes.     

Versatile Solidified Nanofibrous Cellulose-Containing Media for Growth of Extremophiles

Solidified media that employ a porous matrix of nanofibrous cellulose are described. The physicochemical stability of the porous structure allows the development of solidified media that can support the growth of extremophiles, such as acidophilic Acidiphilium, alkaliphilic Bacillus, thermophilic Geobacillus and Thermus, alkalithermophilic Bacillus, and acidothermophilic Sulfolobus microbes. The cellulose-supported media have several advantages over agar- and gellan gum-derived media, including versatility and stability.Solidified media using agar as a solidifying agent are indispensable in microbiology. For solid cultures of mesophilic microorganisms, agar is an ideal solidifying agent and has been used essentially unchanged since it was first introduced in the late 19th century (2, 15). However, the situation is very different when it comes to culturing extremophiles on solidified media. For example, agar media are not suitable for culturing thermophiles and hyperthermophiles because the solidification of agar is thermoreversible at around 50 to 60°C (21), and the media are unstable at temperatures much above 70°C for extended periods (1). Culturing extremophiles on solidified media under acidic or alkaline conditions presents a similar problem of instability.We reported previously the use of porous plates made of nanofibrous cellulose for microbial culture (4). Detailed accounts of the preparation procedure, fine structure of the cellulose plate, and its application to culturing representative mesophilic microorganisms (Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae) appeared in that paper.Interestingly, the porous structure of the cellulose plate has a structural robustness comparable to that of highly crystalline cellulose (6-8) despite its seemingly fragile structure (4). In situ optical microscopic observation in hot and compressed water (5, 18) revealed that the cellulose plate remained unchanged in water up to 260°C at 25 MPa (4, 7). This finding suggested that the cellulose plate could be used, in principle, as a versatile platform for developing solidified media that support the growth of extremophilic microbes under a wide range of extreme culture conditions. A proof-of-concept experiment was done by successfully culturing Thermus thermophilus on the cellulose plate at 80°C (4), but its true potential, especially its versatility, still remained to be corroborated.In this paper, we show that a wide variety of extremophiles, including an acidophile, an alkaliphile, thermophiles, an acidothermophile, and an alkalithermophile, can be cultured on the cellulose plates. Another advantage of the cellulose-supported media, that no solidifying aid is needed for solidification regardless of culture conditions, is also discussed.     

L-Galactono-{gamma}-Lactone Dehydrogenase: Partial Characterization, Induction of Activity and Role in the Synthesis of Ascorbic Acid in Wounded White Potato Tuber Tissue

The total initial vitamin C content (23 mg%) of sliced whitepotato tuber tissue incubated at 20°C decreased slightlyduring the first 12 h and increased thereafter by 80% duringa 48-h incubation. The ascorbic acid (AsA) content (21 mg%)decreased slightly during the first 12 h and then increasedby 40% within 36 h and slowly decreased thereafter. The levelof dehydroascorbic acid (DHA) was low initially, being only2 mg% at 36 h it started to increase and reached 10 mg% at 48h. The activty of L-galactono-     

Thioesterase activity and subcellular localization of acylprotein thioesterase 1/lysophospholipase 1

Acylprotein thioesterase 1 (APT1), also known as lysophospholipase 1, is an important enzyme responsible for depalmitoylation of palmitoyl proteins. To clarify the substrate selectivity and the intracellular function of APT1, we performed kinetic analyses and competition assays using a recombinant human APT1 (hAPT1) and investigated the subcellular localization. For this purpose, an assay for thioesterase activity against a synthetic palmitoyl peptide using liquid chromatography/mass spectrometry was established. The thioesterase activity of hAPT1 was most active at neutral pH, and did not require Ca²⁺ for its maximum activity. The K_M values for thioesterase and lysophospholipase (against lysophosphatidylcholine) activities were 3.49 and 27.3 μM, and the V_max values were 27.3 and 1.62 μmol/min/mg, respectively. Thus, hAPT1 revealed much higher thioesterase activity than lysophospholipase activity. One activity was competitively inhibited by another substrate in the presence of both substrates. Immunocytochemical and Western blot analyses revealed that endogenous and overexpressed hAPT1 were mainly localized in the cytosol, while some signals were detected in the plasma membrane, the nuclear membrane and ER in HEK293 cells. These results suggest that eliminating palmitoylated proteins and lysophospholipids from cytosol is one of the functions of hAPT1.     

Detection of the Leghemoglobin Gene on Two Chromosomes of Phaseolus vulgaris by in situ PCR Linked-Fluorescent in situ Hybridization (FISH)

The leghemoglobin (Lb) gene on the metaphase chromosomes ofPhaseolus vulgaris was amplified by in situ PCR. The amplifiedLb gene could be detected on two chromosomes by fluorescentin situ hybridization (FISH) using the short Lb gene probe. (Received January 9, 1998; Accepted April 30, 1998)