[Function of nitric oxide in initiating production of lignin degrading peroxidases by Phanerochaete chrysosporium].

OBJECTIVE: By analyzing the function and mechanism of nitric oxide in initiating producing lignin peroxidases by phanerochaete chrysosporium, we studied the regulation mechanism triggering the secondary metabolism of white-rot fungi. METHODS: Mutant (pcR5305) and wild-type (pc530) strains of phanerochaete chrysosporium were respectively cultured under both the conditions of nitrogen limitation and nitrogen sufficiency. To compare their lignin peroxidases (LiP)-production and nitric oxide(NO)-production kinetics and their different influences on producing LiP after the NO donor Sodium Nitroprusside (SNP) and scavenger cPTIO were respectively added to the nitrogen limitation or sufficiency culture medium to show the function and mechanism of nitric oxide in initiating production of lignin peroxidases by white-rot fungi. RESULTS: Both strains produced nitric oxide (NO) under the two opposite nutritional conditions, but the levels of NO produced were related with the type of strain and the nutritional conditions. Strain pc530 produced NO requiring nutrition depletion and producing of NO was strongly delayed and reduced when it was cultured under nitrogen sufficiency condition. On the contrary, pcR5305 did not require nitrogen depletion to trigger and the levels of NO were higher than that of pc530. The results indicate that LiP content had positive correlation with NO value except the occurrence time of LiP peak value was later than that of NO. The ability of producing LiP was promoted after the NO donor SNP added, but SNP affected more on pc530 than pcR5305 in promoting producing LiP. 15mM cPTIO would greatly repress producing LiP, but could not completely restrain the synthesis of LiP for both strains. CONCLUSION: By producing NO, Phanerochaete chrysosporium triggers LiP synthesis. However, the evidences do not indicate that NO participates or effect directly in LiP synthesis. It is more likely that NO is reacting as an upstream signal molecule. Besides NO, there are other signal molecules that have a positive effect on NO levels also involving in the regulation producing LiP. The mechanism of the resistance to nutritional repression of pcR5305 in synthesizing lignin degrading peroxidases may be the answer to the different NO production mechanism of pcR5305 from pc530.

SENP1 deficiency promotes ER stress-induced apoptosis by increasing XBP1 SUMOylation.

The transcription factor X box-binding protein 1 (XBP1) is a key component of the endoplasmic reticulum (ER) stress response. Recently, it has been reported that the spliced XBP1 (XBP1s), an activated XBP1 during ER stress, can be SUMOylated. Here, we identify Sentrin/SUMO-specific protease 1 (SENP1) as a specific de-SUMOylation protease for XBP1. SENP1 can increase the transcriptional activity of XBP1. In Senp1 (-/-) cells, the SUMOylated XBP1 is accumulated, and the expression of XBP1 target genes is downregulated in response to ER stress. Moreover, SENP1 deficiency significantly increases ER stress-induced apoptosis through accumulating XBP1 SUMOylation. These results reveal an essential function of SENP1 in ER stress response through regulating XBP1 SUMOylation.

SUMO-specific protease 2 is essential for suppression of polycomb group protein-mediated gene silencing during embryonic development.

SUMO-specific protease 2 (SENP2) has a broad de-SUMOylation activity in vitro. However, the biological function of SENP2 is largely unknown. Here, we show that deletion of SENP2 gene in mouse causes defects in the embryonic heart and reduces the expression of Gata4 and Gata6, which are essential for cardiac development. SENP2 regulates transcription of Gata4 and Gata6 mainly through alteration of occupancy of Pc2/CBX4, a polycomb repressive complex 1 (PRC1) subunit, on its promoters. We demonstrate that Pc2/CBX4 is a target of SENP2 in vivo and that SUMOylation is essential for Pc2/CBX4-mediated PRC1 recruitment to methylated histone 3 at K27 (H3K27me3). In SENP2 null embryos, SUMOylated Pc2/CBX4 accumulates and Pc2/CBX4 occupancy on the promoters of PcG target genes is markedly increased, leading to repression of Gata4 and Gata6 transcription. Our results reveal a critical role for de-SUMOylation in the regulation of PcG target gene expression.

Autografts and xenografts of skin fibroblasts delivering BMP-2 effectively promote orthotopic and ectopic osteogenesis.

Typical ex vivo bone morphogenetic protein-2 (BMP-2) gene therapy for localized bone formation usually utilizes bone marrow stromal cells as gene delivering cells. Skin fibroblasts, which are abundant and easily obtained, have potential advantages for autologous transplantation, but this application has not been adequately investigated. The purpose of this study was to determine the osteogenetic potential of fibroblasts delivering human BMP-2 (hBMP-2) gene by a retroviral system. The phenotypes of osteogenesis in human dermal fibroblasts transduced with hBMP-2 were determined in vitro. Ectopic osteogenesis was evaluated following the injection of these cell xenografts into muscles of null mice, and the potential for orthotopic bone regeneration was evaluated from syngrafts and autografts of rat dermal fibroblasts in rat calvaria defects. The activity of alkaline phosphatase and expression of osteocalcin in fibroblasts transduced with hBMP-2 were increased, and ectopic osteogenesis could be detected in muscles from null mice. The syngrafts and autografts of rat dermal fibroblasts transduced with hBMP-2 gene significantly promoted bone repair and partially healed the calvarial defects. Syngrafts and autografts of rat fibroblasts transduced with hBMP-2 gene had greater average areas exhibiting an osteogenic response compared with the control. The success of bone regeneration in calvaria defects induced by the autologous hBMP-2-modified skin fibroblasts provides evidence that fibroblasts could be effectively used in ex vivo gene therapy for local bone repair.