Cellular abscisic acid (ABA) concentration is determined by both de novo biosynthesis and recycling via b-glucosidase(s). However, which rice b-glucosidase(s) are involved in this process remains unknown. Here, we report on a chloroplastic b-glucosidase isoenzyme, Os3BGlu6, that functions in ABA recycling in rice.Disruption of Os3BGlu6 in rice resulted in dwarfism, lower ABA content in leaves, droughtsensitivity, lower photosynthesis rate and higher intercellular CO 2 concentration. Os3BGlu6 could hydrolyze ABA-GE to ABA in vitro. The reversion and overexpression rice lines restored or increased the drought tolerance as shown by the higher b-glucosidase activity, ABA concentrations and expressions of ABA-and drought-responsive genes. Drought induced Os3BGlu6 to form dimers, and the degree of polymerization correlated well with the increase in cellular ABA concentrations and drought tolerance in rice.Os3BGlu6 was responsive to drought and ABA treatments, and the protein was localized to the chloroplast. Disruption of Os3BGlu6 resulted in the increased stomatal density and impaired stomatal movement. Transcriptomics revealed that disruption of Os3BGlu6 resulted in chloroplastic oxidative stress and lowered Rubisco activity even under normal conditions. Taken together, these results suggest that chloroplastically localized Os3BGlu6 significantly affects cellular ABA pools, thereby affecting drought tolerance and photosynthesis in rice.

Heat-shock protein 70 (HSP70) is ubiquitously found in a variety of organisms and plays an important role in cytoprotection, environmental monitoring, and disease resistance. In this study, the full-length complementary DNA (cDNA) of hsp70 from planarian Polycelis sp. was first cloned using rapid amplification of cDNA ends (RACE). The expression levels of Pyhsp70 were analyzed in the presence of various stressors by real-time PCR, and its temporal-spatial expression patterns were also examined in both intact and regenerative animals by whole-mount in situ hybridization. The results show that (1) the deduced amino acid sequence of Pyhsp70 includes three typical HSP70 family signature motifs and is highly conserved during evolution; (2) Pyhsp70 expression is induced by prolonged starvation, tissue damage, and ionic liquid but inhibited by high or low temperatures; and (3) Pyhsp70 mRNA is mainly expressed in the head peripheral region and in the regenerating blastema during regeneration. These results suggest that the highly expressed Pyhsp70 gene may contribute to enhance cytoprotection and tolerance against stress-induced molecular damage, and the migration of neoblasts to the wound, which might also be involved in the proliferation and differentiation of neoblasts. Our work provides basic data for the study of stress responses and regenerative mechanism in freshwater planarians.

Planarians provide the ideal model for studying eye development, with their simple eye structure and exceptionally rapid regeneration. Here, we observed the eye morphogenesis, photophobic behavior, spectral sensitivity and expression pattern of Djopsin in the freshwater planarian Dugesia japonica. The results showed that: (i) Djopsin encoding the putative protein belonged to the rhabdomeric opsins group and displayed high conservation during animal evolution; (ii) planarians displayed diverse photophobic response to different visible wavelengths and were more sensitive to light blue (495 nm) and yellow (635 nm); (iii) the morphogenesis and functional recovery of eyes were related to the expression pattern of Djopsin during head regeneration; and (iv) Djopsin gene plays a major role in functional recovery during eye regeneration and visual system maintenance in adult planarians.

Identification and evolution of salt tolerant genes are crucial steps in developing salt tolerant crops or microorganisms using biotechnology. Ds-26-16, a salt tolerant gene that was isolated from Dunaliella salina, encodes a transcription factor that can confer salt tolerance to a number of organisms including Escherichia coli (E. coli), Haematococcus pluvialis and tobacco. To further improve its salt tolerance, a random mutagenesis library was constructed using deoxyinosine triphosphate-mediated error-prone PCR technology, and then screened using an E. coli expression system that is based on its broad-spectrum salt tolerance. Seven variants with enhanced salt tolerance were obtained. Variant EP-5 that contained mutation S32P showed the most improvement with the E. coli transformant enduring salt concentrations up to 1.54 M, in comparison with 1.03 M for the wild type gene. Besides, Ds-26-16 and EP-5 also conferred E. coli transformant tolerance to freezing, cold, heat, Cu2+ and alkaline. Homology modeling revealed that mutation S32P in EP-5 caused the conformational change of N- and C-terminal α-helixes. Expression of Ds-26-16 and EP-5 maintained normal cellular morphology, increased the intracellular antioxidant enzymatic activity, reduced malondialdehyde content, and stimulated Nitric Oxide synthesis, thus enhancing salt tolerance to E. coli transformants.

Planarians, the representatives of an ancient bilaterian group with complex reproductive system and high regenerative capabilities, are model system suitable for studying the basic molecular requirements for the development of the reproductive system. To further explore the morphological changes of the gonads during desexualization and the molecular events of the genes controlling the reproductive system development in planarians, we have investigated the histological changes of ovary and testis by paraffin section and the expression patterns of reproductive-related genes by the quantitative real-time PCR in Dugesiajaponica Ichikawa & Kawakatsu, 1964, upon starvation. The four genes, Djprps, DjvlgA, DjvlgB and Djnos, have been selected. The research results show that the degradation of ovary changes from outside layer to inside, and the testis changes are opposite; the reproductive capacity of the planarians starts to be damaged from the 17th to 25th days and to disappear completely from the 26th to 37th days during starvation. The expression patterns of the four genes exhibit the obvious dynamic variations during their desexualization, which indicates that these genes might be involved in gonad development.