Arabidopsis 3ß-Hydroxysteroid Dehydrogenases/C4-Decarboxylases Are Essential for the Pollen and Embryonic Development.

The biosynthesis of C27-29 sterols from their C30 precursor squalene involves C24-alkylation and the removal of three methyl groups, including two at the C4 position. The two C4 demethylation reactions require a bifunctional enzyme known as 3ß-hydroxysteroid dehydrogenase/C4-decarboxylase (3ßHSD/D), which removes an oxidized methyl (carboxylic) group at C4 while simultaneously catalyzing the 3ß-hydroxyl→3-keto oxidation. Its loss-of-function mutations cause ergosterol-dependent growth in yeast and congenital hemidysplasia with ichthyosiform erythroderma and limb defect (CHILD) syndrome in humans. Although plant 3ßHSD/D enzymes were well studied enzymatically, their developmental functions remain unknown. Here we employed a CRISPR/Cas9-based genome-editing approach to generate knockout mutants for two Arabidopsis 3ßHSD/D genes, HSD1 and HSD2, and discovered the male gametophytic lethality for the hsd1 hsd2 double mutation. Pollen-specific expression of HSD2 in the heterozygous hsd1 hsd2/+ mutant not only rescued the pollen lethality but also revealed the critical roles of the two HSD genes in embryogenesis. Our study thus demonstrated the essential functions of the two Arabidopsis 3ßHSD/D genes in male gametogenesis and embryogenesis.

Plastid genome sequencing, identification of nuclear SNP markers, and quality assessment of medicinal rhizomatous herb Polygonatum odoratum (Asparagaceae) cultivars.

Polygonatum odoratum (Mill.) Druce (Asparagaceae, Asparagales) is a widely cultivated medicinal herb in China. However, this useful herb is understudied despite being known as a medicinal resource with top grade medical and edible properties since long. In this study, P. odoratum and four cultivars were investigated. The variations in morphological characteristics and vegetative phases of each cultivar were observed. For genetic aspect, the plastid genome of P. odoratum varies in length from 154,569 bp to 155,491 bp, containing a large single-copy region of 83,486-84,459 bp, a small single-copy region of 18,292-18,471 bp, and two inverted repeats of 26,302-26,370 bp. A total of 131 genes were predicted, including 85 protein-coding, 38 tRNA, and eight rRNA genes. Genome comparisons revealed a slight variation in the sequence across the five accessions, but two highly variable regions (trnC-petN and rpl32-trnL) were detected when comparing the four different cultivars. For the RAD-seq markers, a total of 33.64 Gb of clean data, with an average value of 1.08 Gb per sample, were analyzed for the presence of single nucleotide polymorphisms (SNPs). Well-resolved phylogenies of the P. odoratum cultivars are constructed; the nonmonophyletic relationship in the plastome-based phylogenetic trees, yet monophyletic form in the RAD-based linkage map suggested possibility of hybrid cultivar for P. odoratum "Dazhu" (GDDZ), which was further supported by morphological observations. Quality assessment based on the standards of the Chinese Pharmacopoeia on Polygonati Odorati Rhizoma (POR) on the four cultivars used in this study recorded that PORs from P. odoratum "Zhongzhu" (GDZZ) met the minimum criteria for the acceptance as raw material for medicinal drug production. This study has provided insights on the morphological variations, genetic background, and medicinal qualities of P. odoratum cultivars that could be explored for future genetic improvement as well as breeding programs of P. odoratum for POR production.

Irisin promotes cementoblast differentiation via p38 MAPK pathway.

OBJECTIVE: Irisin is a newly identified exercise-induced myokine which can affect glucose metabolism and cortical bone mass and strength. However, the influence of irisin on cementoblasts remains largely unknown. MATERIAL AND METHODS: An immortalized mouse cementoblast cell line OCCM-30 was used in this study. Cementoblast differentiation markers and PGC-1α in cells cultured with mineral induction medium were evaluated by qRT-PCR. Cementoblast mineralization was evaluated by alizarin red staining. Differentiation markers and the activity of p38 MAPK pathway under irisin stimulation were assessed by qRT-PCR or Western blot analysis. p38 MAPK pathway inhibitor SB203580 or p38 siRNA was used to further identify the regulatory mechanism. Cell proliferation treated with irisin was examined by CCK-8 method. RESULTS: The expression of Runx2, osterix, ALP, and PGC-1α was up-regulated consistently under mineral induction. The formation of mineralized nodules was increased by irisin. Runx2, osterix, ALP, and osteocalcin were obviously up-regulated under irisin stimulation as well as the activity of p38 MAPK pathway. When pretreated with SB203580 or p38 siRNA before irisin stimulation, the irisin-induced differentiation was distinctly suppressed. OCCM-30 cell proliferation was enhanced when treated with high-dose irisin for long time. CONCLUSION: Irisin can promote the differentiation of cementoblasts via p38 MAPK pathway.