151 research outputs found
Tomato plants transgenic for an Arabidopsis thaliana cystein proteinase inhibitor (Atcys) impair the life cycle of Helicoverpa armigera (Hüb.)
Atcys tomato (Lycopersicon esculentum Mill.) transgenic plants, expressing a cystein proteinase inhibition level double than the untransformed control (Speranza et al. in press), were used for in vivo assays with H. armigera larvae. This insect pest, extremely polyphagous, has recently caused severe damages to the outdoor tomato crop due to the dropping of infested young fruits and to fruit rotting because of the larval galleries.
Plants of the cv. Riogrande (RIG) and of the corresponding Atcys homozygous transgenic line (BG-106) were grown in greenhouse and leaves utilized for feeding H. armigera larvae, reared for four days with artificial diet. The recorded data were larval weight (every two days until the cocoon stage), cocoon sex and morphometric traits, number of adults emerged from the cocoon, number of layed and hatched eggs. The mean weight was generally higher when larvae were fed with BG-106 leaves. By subdividing in three periods the larval life, no difference in mortality was observed between larvae reared with control (RIG) and with BG-106 leaves. The percentage of adults emerged from the cocoon was 81% and 76% for the control and BG-106 respectively. The sex ratio (males/females) was in favour of the female sex both for the RIG (0.87) and BG-106 (0.73) cocoons. On average, the fertility (number of layed eggs) of the BG-106 fed females was 33% lower than the control. By considering the percentage of hatched eggs (emerged larvae), the value obtained was 6.8% for BG-106 against 11% for RIG.
According to these data, in Atcys transgenic tomato (BG-106), a level of cystein proteinase inhibition double than the untransformed control, is sufficient to negatively influence the H. armigera biological cycle, even if the weight of the larvae fed with the BG-106 leaves is on average higher than the control (RIG). The last datum is in agreement with similar experiments reported in literature where the effect of proteinase inhibitors is tested in different host-pest systems
Mass spectrometric analysis of the HMGY protein from Lewis lung carcinoma. Identification of phosphorylation sites.
The primary structure of the Lewis lung carcinoma protein HMGY belonging to the nuclear group of proteins HMGI (high mobility group I) was determined using electrospray and fast atom bombardment mass spectrometry. It was demonstrated that the sequence of the tumor protein corresponds to the amino acid sequence derived from the cDNA from cultured cells and that the N-terminal serine residue is N-acetylated. Moreover, the two high performance liquid chromatography-purified forms Y1 and Y2 of the protein HMGY were shown to differ at the level of serine phosphorylation, since they contain three phosphate and two phosphate groups, respectively, in the C-terminal region. No other modification was detected in the remaining part of the molecule
Identification of proteinaceous binders in paintings: A targeted proteomic approach for cultural heritage
Abstract Identification of proteins in paintings and polychrome objects is a challenge, which requires the development of tailored analytical approaches. In the present study, a targeted proteomics approach was developed for discriminating among the three most common proteinaceous materials used as paint binders, i.e. milk, egg, and animal glue. In this study a specific database of peptides was created based on tandem MS analyses of tryptic digests of several paint samples collected from a variety of art objects of different ages and conservation conditions. Specific peptide markers of each protein were then selected and monitored by LC-MSMS in Multiple Reaction Monitoring (MRM) ion mode, together with their specific precursor ion-product ion transitions, as defined by their unique amino acid sequence. The developed method enabled a sensitive and reliable detection of the target peptides in a selection of case studies, leading to the unambiguous identification of the proteins used as paint binders. The method showed greatly increased sensitivity compared to currently available strategies
Genome-wide mapping of 8-oxo-7,8-dihydro-2'-deoxyguanosine reveals accumulation of oxidatively-generated damage at DNA replication origins within transcribed long genes of mammalian cells
8-Oxo-7,8-dihydro-2-deoxyguanosine (8-oxodG) is
one of the major DNA modifications and a potent
pre-mutagenic lesion prone to mispair with 2-
deoxyadenosine (dA). Several thousand residues of
8-oxodG are constitutively generated in the genome
of mammalian cells, but their genomic distribution
has not yet been fully characterized. Here, by
using OxiDIP-Seq, a highly sensitive methodology
that uses immuno-precipitation with efficient anti–
8-oxodG antibodies combined with high-throughput
sequencing, we report the genome-wide distribution
of 8-oxodG in human non-tumorigenic epithelial
breast cells (MCF10A), and mouse embryonic
fibroblasts (MEFs). OxiDIP-Seq revealed sites of 8-
oxodG accumulation overlapping with H2AX ChIPSeq
signals within the gene body of transcribed long
genes, particularly at the DNA replication origins
contained therein. We propose that the presence of
persistent single-stranded DNA, as a consequence
of transcription-replication clashes at these sites, determines
local vulnerability to DNA oxidation and/or
its slow repair. This oxidatively-generated damage,
likely in combination with other kinds of lesion, might
contribute to the formation of DNA double strand
breaks and activation of DNA damage response
The TRILL project: increasing the technological readiness of Laue lenses
Hard X-/soft Gamma-ray astronomy (> 100 keV) is a crucial field for the study
of important astrophysical phenomena such as the 511 keV positron annihilation
line in the Galactic center region and its origin, gamma-ray bursts, soft
gamma-ray repeaters, nuclear lines from SN explosions and more. However,
several key questions in this field require sensitivity and angular resolution
that are hardly achievable with present technology. A new generation of
instruments suitable to focus hard X-/soft Gamma-rays is necessary to overcome
the technological limitations of current direct-viewing telescopes. One
solution is using Laue lenses based on Bragg's diffraction in a transmission
configuration. To date, this technology is in an advanced stage of development
and further efforts are being made in order to significantly increase its
technology readiness level (TRL). To this end, massive production of suitable
crystals is required, as well as an improvement of the capability of their
alignment. Such a technological improvement could be exploited in stratospheric
balloon experiments and, ultimately, in space missions with a telescope of
about 20 m focal length, capable of focusing over a broad energy pass-band. We
present the latest technological developments of the TRILL (Technological
Readiness Increase for Laue Lenses) project, supported by ASI, devoted to the
advancement of the technological readiness of Laue lenses. We show the method
we developed for preparing suitable bent Germanium and Silicon crystals and the
latest advancements in crystals alignment technology.Comment: arXiv admin note: text overlap with arXiv:2211.1688
Wilson Disease Protein ATP7B Utilizes Lysosomal Exocytosis to Maintain Copper Homeostasis
SummaryCopper is an essential yet toxic metal and its overload causes Wilson disease, a disorder due to mutations in copper transporter ATP7B. To remove excess copper into the bile, ATP7B traffics toward canalicular area of hepatocytes. However, the trafficking mechanisms of ATP7B remain elusive. Here, we show that, in response to elevated copper, ATP7B moves from the Golgi to lysosomes and imports metal into their lumen. ATP7B enables lysosomes to undergo exocytosis through the interaction with p62 subunit of dynactin that allows lysosome translocation toward the canalicular pole of hepatocytes. Activation of lysosomal exocytosis stimulates copper clearance from the hepatocytes and rescues the most frequent Wilson-disease-causing ATP7B mutant to the appropriate functional site. Our findings indicate that lysosomes serve as an important intermediate in ATP7B trafficking, whereas lysosomal exocytosis operates as an integral process in copper excretion and hence can be targeted for therapeutic approaches to combat Wilson disease
The gene of an archaeal α-l-fucosidase is expressed by translational frameshifting
The standard rules of genetic translational decoding are altered in specific genes by different events that are globally termed recoding. In Archaea recoding has been unequivocally determined so far only for termination codon readthrough events. We study here the mechanism of expression of a gene encoding for a α-l-fucosidase from the archaeon Sulfolobus solfataricus (fucA1), which is split in two open reading frames separated by a −1 frameshifting. The expression in Escherichia coli of the wild-type split gene led to the production by frameshifting of full-length polypeptides with an efficiency of 5%. Mutations in the regulatory site where the shift takes place demonstrate that the expression in vivo occurs in a programmed way. Further, we identify a full-length product of fucA1 in S.solfataricus extracts, which translate this gene in vitro by following programmed −1 frameshifting. This is the first experimental demonstration that this kind of recoding is present in Archaea
Different Factors Affecting Human ANP Amyloid Aggregation and Their Implications in Congestive Heart Failure
Atrial Natriuretic Peptide (ANP)-containing amyloid is frequently found in the elderly heart. No data exist regarding ANP aggregation process and its link to pathologies. Our aims were: i) to experimentally prove the presumptive association of Congestive Heart Failure (CHF) and Isolated Atrial Amyloidosis (IAA); ii) to characterize ANP aggregation, thereby elucidating IAA implication in the CHF pathogenesis.A significant prevalence (85\%) of IAA was immunohistochemically proven ex vivo in biopsies from CHF patients. We investigated in vitro (using Congo Red, Thioflavin T, SDS-PAGE, transmission electron microscopy, infrared spectroscopy) ANP fibrillogenesis, starting from α-ANP as well as the ability of dimeric β-ANP to promote amyloid formation. Different conditions were adopted, including those reproducing β-ANP prevalence in CHF. Our results defined the uncommon rapidity of α-ANP self-assembly at acidic pH supporting the hypothesis that such aggregates constitute the onset of a fibrillization process subsequently proceeding at physiological pH. Interestingly, CHF-like conditions induced the production of the most stable and time-resistant ANP fibrils suggesting that CHF affected people may be prone to develop IAA.We established a link between IAA and CHF by ex vivo examination and assessed that β-ANP is, in vitro, the seed of ANP fibrils. Our results indicate that β-ANP plays a crucial role in ANP amyloid deposition under physiopathological CHF conditions. Overall, our findings indicate that early IAA-related ANP deposition may occur in CHF and suggest that these latter patients should be monitored for the development of cardiac amyloidosis
Impact of different chemotherapy regimens on intestinal mucosal injury assessed with bedside ultrasound: a study in 213 AML patients
IntroductionNeutropenic enterocolitis (NEC) is a life-threatening complication reported in patients with acute myeloid leukemia (AML) following chemotherapy (CHT). Intensive induction and consolidation CHT may damage intestinal mucosa leading to a NEC episode (NECe). NEC reported mortality may be up to 30-60%. Early US-guided bed-side diagnosis and prompt treatment may substantially improve the survival. An emerging worldwide concern is the intestinal colonization by multi-drug-resistant bacteria especially when patients are exposed to chemotherapy regimens potentially correlated to mucosal damage. MethodsIn our study we prospectively enrolled all AML patients admitted in our leukemia unit to receive intensive induction and consolidation chemotherapy and experiencing chemotherapy-induced-neutropenia (CHTN). Results and discussionOverall, we enrolled N=213 patients from 2007 to March 2023. We recorded N=465 CHTN, and N=42 NECe (9.0% incidence). The aim of our study was to assess which chemotherapy regimens are more associated with NEC. We found that ALM1310, followed by 7 + 3 (daunorubicin), 7 + 3 (idarubicin), 5 + 3 + 3 (cytarabine, etoposide, idarubicin), and AML1310 (consolidation) were associated with a statistically higher incidence of NEC. We did not detect NEC episodes in patients treated with CPX-351, 5 + 2 (cytarabine, idarubicine), and high-dose cytarabine. Thus, we found that cytarabine could determine mucosal damage when associated with an anthracycline but not if delivered either alone or as dual-drug liposomal encapsulation of daunorubicin/cytarabine. We also describe NEC mortality, symptoms at diagnosis, intestinal sites involvement, and prognostic significance of bowel wall thickening
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