Targeted Quantification of the Glycated Peptides of Human Serum Albumin

Vannuruswamy, Garikapati; Korwar, Arvind M.; Jagadeeshaprasad, Mashanipalya G.; Kulkarni, Mahesh J.

doi:10.1007/978-1-4939-7057-5_28

Cited by 2 publications

(2 citation statements)

References 9 publications

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“…Protein samples were subjected to in-solution tryptic digestion followed by mass spectrometry (MS) analysis as described earlier with minor modifications [87]. In brief, proteins (20 µg) were diluted with 50 mM ammonium bicarbonate buffer containing 0.1% RapiGest (Waters), followed by incubation at 80°C for 15 min.…”

Section: In-solution Tryptic Digestionmentioning

confidence: 99%

“…Pellets of isolated EV fractions were washed twice with 50 mM ammonium bicarbonate buffer using Amicon Ultra 3 kDa cut-off centrifugal filters (Merck Millipore Ltd), and total protein concentration was estimated using Quick Start Bradford Protein Assay (Bio-Rad Laboratories). Protein samples were subjected to in-solution tryptic digestion followed by mass spectrometry (MS) analysis as described earlier with minor modifications (Vannuruswamy et al 2017). In brief, proteins (20 µg) were diluted with 50 mM ammonium bicarbonate buffer containing 0.1% RapiGest (Waters Corporation), followed by incubation at 80°C for 15 min.…”

Section: In-solution Tryptic Digestionmentioning

confidence: 99%

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Host-induced gene silencing involves Arabidopsis ESCRT-III pathway for the transfer of dsRNA-derived siRNA

Schlemmer

Weipert

Barth

et al. 2020

Preprint

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Small (s)RNA molecules are crucial factors in the communication between hosts and their interacting pathogens, where they function as effectors that can modulate both host defense and microbial virulence/pathogenicity through a mechanism termed cross-kingdom RNA interference (ckRNAi). Consistent with this recent knowledge, sRNAs and their doublestranded (ds)RNA precursors have been adopted to control diseases in crop plants through transgenic expression (host-induced gene silencing, HIGS) or exogenous application (sprayinduced gene silencing, SIGS). While these strategies proved to be effective, the mechanism of RNA transfer at the plant -pathogen interface is widely unresolved. Here we show that extracellular vesicles (EVs) purified from Arabidopsis (Arabidopsis thaliana) leaf extracts and apoplastic fluids contain transgene-derived sRNAs. EVs from plants expressing CYP3RNA, a 791 nt long dsRNA, which was originally designed to target the three CYP51 genes of the fungal pathogen Fusarium graminearum, contain CYP3RNA-derived small interfering (si)RNAs as shown by RNA sequencing (RNA-seq) analysis. Notably, the EVs cargo retained the same CYP3RNA-derived siRNA profile as the respective leaf extracts, suggesting that there was no selective uptake of specific artificial sRNAs into EVs. In addition, mutants of the ESCRT-III complex were impaired in HIGS further indicating that endosomal vesicle trafficking supports transfer of transgene-derived siRNAs between donor host cells and recipient fungal cells.Further supporting the relevance of EV-mediated transport of sRNA, we demonstrate that HIGS plants, expressing a 100 nt dsRNA-target-sequence identified via EV-sRNA-seq of CYP3RNA Arabidopsis, confers strong resistance to F. graminearum. Together, these findings support the view that EVs are key mediators in the transport of HIGS-related sRNAs to reduce the virulence of interacting fungal pathogens during host-pathogen interaction.(MVBs) fuse with the PM [33][34][35][36][37]. In mammals, exosomes mediate intercellular communication by shuttling proteins, lipids and RNAs, where RNA molecules remain functional after delivery and can elicit effects in the recipient cell [38][39][40].Based on this knowledge, we speculated that HIGS-and SIGS-related RNAs also are loaded into plant vesicles and transferred by EVs that cross the plant-fungus interface. It was reasoned already more than 50 years ago that a fusion of plant MVBs with the PM may result in the release of small vesicles into the extracellular space [41]. Consistent with this early observation, biogenesis of immune-related MVBs [29,30] and the release of their cargo via EVs also is inherent to the plant secretory pathway [27]. Transmission electron microscopy (TEM) revealed proliferation of MVBs next to plant cell wall papillae in response to infection with powdery mildew fungus and the release of para-mural vesicles. The authors proposed that released vesicles might be similar to exosomes in animal cell thus anticipating that exosomes exist in plants [29,30]. Immun...

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Section: In-solution Tryptic Digestionmentioning

confidence: 99%

Section: In-solution Tryptic Digestionmentioning

confidence: 99%

Host-induced gene silencing involves Arabidopsis ESCRT-III pathway for the transfer of dsRNA-derived siRNA

Schlemmer

Weipert

Barth

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

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P301 L, an FTDP-17 Mutant, Exhibits Enhanced Glycation in vitro

Sonawane

Chinnathambi

2020

JAD

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Background: Frontotemporal dementia and parkinsonism-linked to chromosome-17 are a group of diseases with tau mutations leading to primary tauopathies which include progressive supranuclear palsy, corticobasal syndrome, and frontotemporal lobar degeneration. Alzheimer’s disease is a non-primary tauopathy, which displays tau neuropathology of excess tangle formation and accumulation. FTDP-17 mutations are responsible for early onset of AD, which can be attributed to compromised physiological functions due to the mutations. Tau is a microtubule-binding protein that secures the integrity of polymerized microtubules in neuronal cells. It malfunctions owing to various insults and stress conditions-like mutations and post-translational modifications. Objective: In this study, we modified the wild type and tau mutants by methyl glyoxal and thus studied whether glycation can enhance the aggregation of predisposed mutant tau. Methods: Tau glycation was studied by fluorescence assays, SDS-PAGE analysis, conformational evaluation, and transmission electron microscopy. Results: Our study suggests that FTDP-17 mutant P301 L leads to enhanced glycation-induced aggregation as well as advanced glycation end products formation. Glycation forms amorphous aggregates of tau and its mutants without altering its native conformation. Conclusion: The metabolic anomalies and genetic predisposition have found to accelerate tau-mediated neurodegeneration and prove detrimental for the early-onset of Alzheimer’s disease.

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Targeted Quantification of the Glycated Peptides of Human Serum Albumin

Cited by 2 publications

References 9 publications

Host-induced gene silencing involves Arabidopsis ESCRT-III pathway for the transfer of dsRNA-derived siRNA

Host-induced gene silencing involves Arabidopsis ESCRT-III pathway for the transfer of dsRNA-derived siRNA

P301 L, an FTDP-17 Mutant, Exhibits Enhanced Glycation in vitro

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