Sequence polymorphisms in ribosomal RNA genes and variations in chromosomal loci of Oenothera odorata and O. laciniata

Numerous studies on Oenothera species have been investigated for the physiological and ecological characteristics. However, no such an information based on molecular cytogenetic has yet been introduced, in turn, is very essential for identifying sequence polymorphisms of rRNA genes with their loci on mitotic phases for further biological researches. In this study, sequence variations of rRNA genes in Oenothera odorata and O. laciniata were examined to identify informative factors as unique or repeat sequences in intra- and inter-specific variations. Intra-specific variation revealed that the sequences of the rRNA genes including the spacer regions were highly conserved revealing only a few variations. From the inter-specific variation, spacer regions of species were completely different as (1) non-homologous sequences in NTS and (2) different type repeat sequences in ITS 1, 2 and 5.8S rRNA, whereas the remaining coding regions were highly conserved. FISH was carried out on mitotic phases using the 5S rDNA of the analyzed sequences. From the interphase and metaphase chromosomes of the examined species, two loci of 5S rDNA in O. odorata and four loci in O. laciniata were confirmed on the telomeric region of the short arm. Due to the small size and unclear centromere of the chromosomes, karyotype could not be completed. However, we confirmed that the chromosomes are organized by meta- and acrocentric chromosomes and the chromosomes with identified loci were assumed to be paired by the location of loci at the telomeric region on the short arm with relative lengths.     

Variations in enzyme activity in stomach and pancreatic tissue and digesta in piglets around weaning

A study was performed to investigate the effect of weaning at 4 weeks of age on the activity of digestive enzymes in the stomach and pancreatic tissue and in digesta from 3 days prior to weaning to 9 days postweaning in 64 piglets. In stomach tissue the activity of pepsin and gastric lipase was determined. Pepsin activity declined abruptly after weaning but 5 days postweaning the weaning level was regained and in the gastric contents no change in pepsin activity was observed. Weaning did not influence the activity of gastric lipase. The activity of eight enzymes and a cofactor was measured in pancreatic tissue. The effect of weaning on the enzyme activity was highly significant for all enzymes except elastase. The activity of all enzymes remained at the weaning level during day 1–2 postweaning followed by a reduction of the activity. The activity of trypsin, carboxypeptidase A, amylase and lipase exhibited minimum activity 5 days postweaning. Trypsin activity increased to the preweaning level on day 7–9 whereas the activity of the others increased but did not reach the preweaning level. The activity of chymotrypsin, carboxypeptidase B and carboxyl ester hydrolase decreased during the entire experimental period. In digesta no effect of weaning was observed on the activity of amylase and trypsin. The activity of chymotrypsin was reduced after weaning in the proximal third of the small intestine and lipase and carboxyl ester hydrolase activity was reduced in the middle and distal parts of the small intestine after weaning. The present study shows that the activities of the digestive enzymes in the pancreatic tissue are affected by weaning. Even though the pancreatic secretion cannot be judged from these results they show that the enzymes respond differently to weaning. In general the activity of the digestive enzymes in pancreatic tissue is low on day 5 postweaning which in interaction with other factors may increase the risk of developing postweaning diarrhoea.     

SDS Interferes with SaeS Signaling of Staphylococcus aureus Independently of SaePQ

The Staphylococcus aureus regulatory saePQRS system controls the expression of numerous virulence factors, including extracellular adherence protein (Eap), which amongst others facilitates invasion of host cells. The saePQRS operon codes for 4 proteins: the histidine kinase SaeS, the response regulator SaeR, the lipoprotein SaeP and the transmembrane protein SaeQ. S. aureus strain Newman has a single amino acid substitution in the transmembrane domain of SaeS (L18P) which results in constitutive kinase activity. SDS was shown to be one of the signals interfering with SaeS activity leading to inhibition of the sae target gene eap in strains with SaeS^L but causing activation in strains containing SaeS^P. Here, we analyzed the possible involvement of the SaeP protein and saePQ region in SDS-mediated sae/eap expression. We found that SaePQ is not needed for SDS-mediated SaeS signaling. Furthermore, we could show that SaeS activity is closely linked to the expression of Eap and the capacity to invade host cells in a number of clinical isolates. This suggests that SaeS activity might be directly modulated by structurally non-complex environmental signals, as SDS, which possibly altering its kinase/phosphatase activity.     

SESN2/sestrin2 suppresses sepsis by inducing mitophagy and inhibiting NLRP3 activation in macrophages

Proper regulation of mitophagy for mitochondrial homeostasis is important in various inflammatory diseases. However, the precise mechanisms by which mitophagy is activated to regulate inflammatory responses remain largely unknown. The NLRP3 (NLR family, pyrin domain containing 3) inflammasome serves as a platform that triggers the activation of CASP1 (caspase 1) and secretion of proinflammatory cytokines. Here, we demonstrate that SESN2 (sestrin 2), known as stress-inducible protein, suppresses prolonged NLRP3 inflammasome activation by clearance of damaged mitochondria through inducing mitophagy in macrophages. SESN2 plays a dual role in inducing mitophagy in response to inflammasome activation. First, SESN2 induces “mitochondrial priming” by marking mitochondria for recognition by the autophagic machinery. For mitochondrial preparing, SESN2 facilitates the perinuclear-clustering of mitochondria by mediating aggregation of SQSTM1 (sequestosome 1) and its binding to lysine 63 (Lys63)-linked ubiquitins on the mitochondrial surface. Second, SESN2 activates the specific autophagic machinery for degradation of primed mitochondria via an increase of ULK1 (unc-51 like kinase 1) protein levels. Moreover, increased SESN2 expression by extended LPS (lipopolysaccharide) stimulation is mediated by NOS2 (nitric oxide synthase 2, inducible)-mediated NO (nitric oxide) in macrophages. Thus, Sesn2-deficient mice displayed defective mitophagy, which resulted in hyperactivation of inflammasomes and increased mortality in 2 different sepsis models. Our findings define a unique regulatory mechanism of mitophagy activation for immunological homeostasis that protects the host from sepsis.     

Autophagy induced by AXL receptor tyrosine kinase alleviates acute liver injury via inhibition of NLRP3 inflammasome activation in mice

Severe hepatic inflammation is a common cause of acute or chronic liver disease. Macrophages are one of the key mediators which regulate the progress of hepatic inflammation. Increasing evidence shows that the TAM (TYRO3, AXL and MERTK) family of RTKs (receptor tyrosine kinases), which is expressed in macrophages, alleviates inflammatory responses through a negative feedback loop. However, the functional contribution of each TAM family member to the progression of hepatic inflammation remains elusive. In this study, we explore the role of individual TAM family proteins during autophagy induction and evaluate their contribution to hepatic inflammation. Among the TAM family of RTKs, AXL (AXL receptor tyrosine kinase) only induces autophagy in macrophages after interaction with its ligand, GAS6 (growth arrest specific 6). Based on our results, autophosphorylation of 2 tyrosine residues (Tyr815 and Tyr860) in the cytoplasmic domain of AXL in mice is required for autophagy induction and AXL-mediated autophagy induction is dependent on MAPK (mitogen-activated protein kinase)14 activity. Furthermore, induction of AXL-mediated autophagy prevents CASP1 (caspase 1)-dependent IL1B (interleukin 1, β) and IL18 (interleukin 18) maturation by inhibiting NLRP3 (NLR family, pyrin domain containing 3) inflammasome activation. In agreement with these observations, axl^?/? mice show more severe symptoms than do wild-type (Axl^+/+) mice following acute hepatic injury induced by administration of lipopolysaccharide (LPS) or carbon tetrachloride (CCl₄). Hence, GAS6-AXL signaling-mediated autophagy induction in murine macrophages ameliorates hepatic inflammatory responses by inhibiting NLRP3 inflammasome activation.     

Generation of Chinese cabbage resistant to bacterial soft rot by heterologous expression of <Emphasis Type="Italic">Arabidopsis WRKY75</Emphasis>

Soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) is a serious disease in Chinese cabbage (Brassica rapa L. subsp. pekinensis). To reduce the severity of soft rot symptoms in Chinese cabbage, Arabidopsis AtWRKY75 was introduced into Chinese cabbage by Agrobacterium-mediated transformation, which was previously reported to reduce susceptibility to Pcc infection in Arabidopsis. Three independent Chinese cabbage transgenic lines carrying AtWRKY75 were obtained. The growth phenotypes of AtWRKY75 overexpression (OE) lines were normal. Bacterial soft rot symptoms and Pcc growth were reduced in AtWRKY75-OE Chinese cabbage lines compared with WT plants. In contrast, overexpression of AtWRKY75 had no effect on infection with a hemibiotrophic pathogen, Xanthomonas campestris pv. campestris (Xcc) causing black rot disease. These results are consistent with those observed in the transgenic Arabidopsis. We found that AtWRKY75 activated a subset of Chinese cabbage genes related to defense against Pcc infection, such as Meri15B, BrPR4, and BrPDF1.2 (but not BrPGIP2). Moreover, overexpression of AtWRKY75 caused H₂O₂ production and activation of H₂O₂ scavenge enzyme genes, suggesting that H₂O₂ played a role in AtWRKY75-mediated resistance to Pcc. Together, these results demonstrated that AtWRKY75 decreased the severity of Pcc-caused bacterial soft rot and activated a subset of Pcc infection defense-related genes in Chinese cabbage similar to in Arabidopsis. It is suggested that AtWRKY75 is a candidate gene for use in crop improvement, because it results in reduced severity of disease symptoms without concurrent growth abnormalities.