Temperature tolerance and expression of heat shock protein 70 in the toxic dinoflagellate Alexandrium tamarense (Dinophyceae)

The aim of this study was to investigate the relationship between temperature tolerance and geographical origin in a species. Temperature tolerance and heat shock protein 70 (Hsp70) content were investigated in the toxic dinoflagellate Alexandrium tamarense, which was isolated from temperate and tropical areas. Results of treatment at 0–37 8C for 1 h revealed that 15 8C-cultured Japanese A. tamarense could survive treatment at 0–25 8C, whereas 30 and 37 8C treatment reduced the survival rate. Malaysian A. tamarense cultured at 25 8C survived at 30 8C; however, 37 8C and low temperature treatment reduced the survival rate. After acclimation of both strains at 20 8C, they were treated at 0, 4, 30, and 37 8C. The survival rate of Japanese A. tamarense at 30 8C was slightly increased compared to that of 15 8C-cultured cells. Treatment at 37 8C for 1 h showed no difference between acclimated and unacclimated cells of both strains. At 0 and 4 8C treatment, almost all cells of Japanese A. tamarense survived; however, the Malaysian cells were unable to survive. Both strains of A. tamarense, acclimated at 20 8C, were treated at 30 8C; change in the amount of Hsp70 was analyzed. Western blot analysis revealed that the induction of Hsp70 in the Japanese strain occurred more quickly than in the Malaysian strain. These results indicate that Hsp70 of A. tamarense is a heat stress-inducible protein and the response is different between strains

HIPK2 and extrachromosomal histone H2B are separately recruited by Aurora-B for cytokinesis

Cytokinesis, the final phase of cell division, is necessary to form two distinct daughter cells with correct distribution of genomic and cytoplasmic materials. Its failure provokes genetically unstable states, such as tetraploidization and polyploidization, which can contribute to tumorigenesis. Aurora-B kinase controls multiple cytokinetic events, from chromosome condensation to abscission when the midbody is severed. We have previously shown that HIPK2, a kinase involved in DNA damage response and development, localizes at the midbody and contributes to abscission by phosphorylating extrachromosomal histone H2B at Ser14. Of relevance, HIPK2-defective cells do not phosphorylate H2B and do not successfully complete cytokinesis leading to accumulation of binucleated cells, chromosomal instability, and increased tumorigenicity. However, how HIPK2 and H2B are recruited to the midbody during cytokinesis is still unknown. Here, we show that regardless of their direct (H2B) and indirect (HIPK2) binding of chromosomal DNA, both H2B and HIPK2 localize at the midbody independently of nucleic acids. Instead, by using mitotic kinase-specific inhibitors in a spatio-temporal regulated manner, we found that Aurora-B kinase activity is required to recruit both HIPK2 and H2B to the midbody. Molecular characterization showed that Aurora-B directly binds and phosphorylates H2B at Ser32 while indirectly recruits HIPK2 through the central spindle components MgcRacGAP and PRC1. Thus, among different cytokinetic functions, Aurora-B separately recruits HIPK2 and H2B to the midbody and these activities contribute to faithful cytokinesis

Molecular structure of the largemouth bass (Micropterus salmoides) Myf5 gene and its effect on skeletal muscle growth

Myogenic Regulatory Factors (MRFs), a family of basic helix-loop-helix (bHLH) transcription factors, play important roles in regulating skeletal muscle development and growth. Myf5, the primary factor of MRFs, initiates myogenesis. Its expression pattern during somitomyogenesis in some fish has been revealed. To further study its effect on fish muscle during postembryonic growth, characterization and function analysis of myf5 cDNA were carried out in largemouth bass. The 1,093 bp cDNA sequence was identified by RT-PCR and 3′RACE, then the ORF of Myf5 cDNA was cloned into the expression vector pcDNA3.1(−)/mycHisB. The recombinant plasmid pcDNA3.1(−)/mycHisB-Myf5 was injected into the dorsal muscle of tilapias. RT-PCR and histochemical results showed that the exogenous gene was transcribed and translated in vivo. Its effect on muscle growth focused on myofiber hypertrophy in white muscle 60 days post injection. This indicated that overexpression of Myf5 can promote myogenesis during the fish muscle postembryonic growth period

B cell-intrinsic TBK1 is essential for germinal center formation during infection and vaccination in mice

The germinal center (GC) is a site where somatic hypermutation and clonal selection are coupled for antibody affinity maturation against infections. However, how GCs are formed and regulated is incompletely understood. Here, we identified an unexpected role of Tank-binding kinase-1 (TBK1) as a crucial B cell-intrinsic factor for GC formation. Using immunization and malaria infection models, we show that TBK1-deficient B cells failed to form GC despite normal Tfh cell differentiation, although some malaria-infected B cell-specific TBK1-deficient mice could survive by GC-independent mechanisms. Mechanistically, TBK1 phosphorylation elevates in B cells during GC differentiation and regulates the balance of IRF4/BCL6 expression by limiting CD40 and BCR activation through noncanonical NF-κB and AKT(T308) signaling. In the absence of TBK1, CD40 and BCR signaling synergistically enhanced IRF4 expression in Pre-GC, leading to BCL6 suppression, and therefore failed to form GCs. As a result, memory B cells generated from TBK1-deficient B cells fail to confer sterile immunity upon reinfection, suggesting that TBK1 determines B cell fate to promote long-lasting humoral immunity