Yasushi Ishihama
Sep 1, 2005; 4:1265-1272
Research

David A. Mitchell
Jun 1, 2006; 47:1118-1127
Thematic Reviews

Abnormal changes of neuronal Tau protein, such as phosphorylation and aggregation, are considered hallmarks of cognitive deficits in Alzheimer's disease. Abnormal phosphorylation is thought to precede aggregation and therefore to promote aggregation, but the nature and extent of phosphorylation remain ill-defined. Tau contains ∼85 potential phosphorylation sites, which can be phosphorylated by various kinases because the unfolded structure of Tau makes them accessible. However, methodological limitations (e.g. in MS of phosphopeptides, or antibodies against phosphoepitopes) led to conflicting results regarding the extent of Tau phosphorylation in cells. Here we present results from a new approach based on native MS of intact Tau expressed in eukaryotic cells (Sf9). The extent of phosphorylation is heterogeneous, up to ∼20 phosphates per molecule distributed over 51 sites. The medium phosphorylated fraction Pm showed overall occupancies of ∼8 Pi (± 5) with a bell-shaped distribution; the highly phosphorylated fraction Ph had 14 Pi (± 6). The distribution of sites was highly asymmetric (with 71% of all P-sites in the C-terminal half of Tau). All sites were on Ser or Thr residues, but none were on Tyr. Other known posttranslational modifications were near or below our detection limit (e.g. acetylation, ubiquitination). These findings suggest that normal cellular Tau shows a remarkably high extent of phosphorylation, whereas other modifications are nearly absent. This implies that abnormal phosphorylations at certain sites may not affect the extent of phosphorylation significantly and do not represent hyperphosphorylation. By implication, the pathological aggregation of Tau is not likely a consequence of high phosphorylation.

Felicia Grasso
Dec 1, 2020; 19:1986-1996
Research

Weixian Deng
Dec 22, 2020; 0:RA120.002411v1-mcp.RA120.002411
Research

Congcong Lu
Nov 17, 2020; 0:R120.002257v1-mcp.R120.002257
Review

Daniel J. Geiszler
Dec 1, 2020; 0:TIR120.002216v1-mcp.TIR120.002216
Technological Innovation and Resources

Jianhua Wang
Nov 4, 2020; 0:RA120.002375v1-mcp.RA120.002375
Research

Natural compounds that can stimulate salivary secretion are of interest in developing treatments for xerostomia, the perception of a dry mouth, that affects between 10 and 30% of the adult and elderly population. Chemesthetic transient receptor potential (TRP) channels are expressed in the surface of the oral mucosa. The TRPV1 agonists capsaicin and piperine have been shown to increase salivary flow when introduced into the oral cavity but the sialogogic properties of other TRP channel agonists have not been investigated. In this study we have determined the influence of different TRP channel agonists on the flow and protein composition of saliva. Mouth rinsing with the TRPV1 agonist nonivamide or menthol, a TRPM8 agonist, increased whole mouth saliva (WMS) flow and total protein secretion compared with unstimulated saliva, the vehicle control mouth rinse or cinnamaldehyde, a TRPA1 agonist. Nonivamide also increased the flow of labial minor gland saliva but parotid saliva flow rate was not increased. The influence of TRP channel agonists on the composition and function of the salivary proteome was investigated using a multi-batch quantitative MS method novel to salivary proteomics. Inter-personal and inter-mouth rinse variation was observed in the secreted proteomes and, using a novel bioinformatics method, inter-day variation was identified with some of the mouth rinses. Significant changes in specific salivary proteins were identified after all mouth rinses. In the case of nonivamide, these changes were attributed to functional shifts in the WMS secreted, primarily the over representation of salivary and nonsalivary cystatins which was confirmed by immunoassay. This study provides new evidence of the impact of TRP channel agonists on the salivary proteome and the stimulation of salivary secretion by a TRPM8 channel agonist, which suggests that TRP channel agonists are potential candidates for developing treatments for sufferers of xerostomia.

Amino acid hydroxylation is a common post-translational modification, which generally regulates protein interactions or adds a functional group that can be further modified. Such hydroxylation is currently considered irreversible, necessitating the degradation and re-synthesis of the entire protein to reset the modification. Here we present evidence that the cellular machinery can reverse FIH-mediated asparagine hydroxylation on intact proteins. These data suggest that asparagine hydroxylation is a flexible and dynamic post-translational modification akin to modifications involved in regulating signaling networks, such as phosphorylation, methylation and ubiquitylation.

Plasmodium, the malaria parasite, undergoes a complex life cycle alternating between a vertebrate host and a mosquito vector of the genus Anopheles. In red blood cells of the vertebrate host, Plasmodium multiplies asexually or differentiates into gamete precursors, the male and female gametocytes, responsible for parasite transmission. Sexual stage maturation occurs in the midgut of the mosquito vector, where male and female gametes egress from the host erythrocytes to fuse and form a zygote. Gamete egress entails the successive rupture of two membranes surrounding the parasite, the parasitophorous vacuole membrane and the erythrocyte plasma membrane. In this study, we used the rodent model parasite Plasmodium berghei to design a label-free quantitative proteomic approach aimed at identifying gender-related proteins differentially released/secreted by purified mature gametocytes when activated to form gametes. We compared the abundance of molecules secreted by wild type gametocytes of both genders with that of a transgenic line defective in male gamete maturation and egress. This enabled us to provide a comprehensive data set of egress-related molecules and their gender specificity. Using specific antibodies, we validated eleven candidate molecules, predicted as either gender-specific or common to both male and female gametocytes. All of them localize to punctuate, vesicle-like structures that relocate to cell periphery upon activation, but only three of them localize to the gametocyte-specific secretory vesicles named osmiophilic bodies. Our results confirm that the egress process involves a tightly coordinated secretory apparatus that includes different types of vesicles and may put the basis for functional studies aimed at designing novel transmission-blocking molecules.

O-GlcNAcylation, the addition of a single N-acetylglucosamine residue to serine and threonine residues of cytoplasmic, nuclear, or mitochondrial proteins, is a widespread regulatory post-translational modification. It is involved in response to nutritional status and stress and its dysregulation is associated with diseases ranging from Alzheimer’s to diabetes.  While the modification was first detected over thirty-five years ago, research into the function of O-GlcNAcylation has accelerated dramatically in the last ten years due to the development of new enrichment and mass spectrometry techniques that facilitate its analysis.  This article summarizes methods for O-GlcNAc enrichment, key mass spectrometry instrumentation advancements, particularly those that allow modification site localization, and software tools that allow analysis of data from O-GlcNAc modified peptides.

We developed a simple and rapid method to enrich protein N-terminal peptides, in which the protease TrypN is first employed to generate protein N-terminal peptides without Lys or Arg and internal peptides with two positive charges at their N-termini, and then the N-terminal peptides with or without N-acetylation are separated from the internal peptides by strong cation exchange chromatography according to a retention model based on the charge/orientation of peptides. This approach was applied to 20 μg of human HEK293T cell lysate proteins to profile the N-terminal proteome. On average, 1,550 acetylated and 200 unmodified protein N-terminal peptides were successfully identified in a single LC/MS/MS run with less than 3% contamination with internal peptides, even when we accepted only canonical protein N-termini registered in the Swiss-Prot database. Since this method involves only two steps, protein digestion and chromatographic separation, without the need for tedious chemical reactions, it should be useful for comprehensive profiling of protein N-termini, including proteoforms with neo-N-termini.

Malaria elimination is still pending on the development of novel tools that rely on a deep understanding of parasite biology. Proteins of all living cells undergo a myriad number of posttranslational modifications (PTMs) that are critical to multifarious life processes. An extensive proteome-wide dissection revealed a fine PTM map of most proteins in both Plasmodium falciparum, the causative agent of severe malaria, and the infected red blood cells. More than two-thirds of proteins of the parasite and its host cell underwent extensive and dynamic modification throughout the erythrocytic developmental stage. PTMs critically modulate the virulence factors involved in the host-parasite interaction and pathogenesis. Furthermore, P. falciparum stabilized the supporting proteins of erythrocyte origin by selective de-modification. Collectively, our multiple omic analyses, apart from having furthered a deep understanding of the systems biology of P. falciparum and malaria pathogenesis, provide a valuable resource for mining new antimalarial targets.

Histone post-translational modifications (PTMs) are one of the main mechanisms of epigenetic regulation. Dysregulation of histone PTMs leads to many human diseases, such as cancer. Due to its high-throughput, accuracy, and flexibility, mass spectrometry (MS) has emerged as a powerful tool in the epigenetic histone modification field, allowing the comprehensive and unbiased analysis of histone PTMs and chromatin-associated factors. Coupled with various techniques from molecular biology, biochemistry, chemical biology and biophysics, MS has been employed to characterize distinct aspects of histone PTMs in the epigenetic regulation of chromatin functions. In this review we will describe advancements in the field of MS that have facilitated the analysis of histone PTMs and chromatin biology.  

Open searching has proven to be an effective strategy for identifying both known and unknown modifications in shotgun proteomics experiments. Rather than being limited to a small set of user-specified modifications, open searches identify peptides with any mass shift that may correspond to a single modification or a combination of several modifications. Here we present PTM-Shepherd, a bioinformatics tool that automates characterization of PTM profiles detected in open searches based on attributes such as amino acid localization, fragmentation spectra similarity, retention time shifts, and relative modification rates. PTM-Shepherd can also perform multi-experiment comparisons for studying changes in modification profiles, e.g. in data generated in different laboratories or under different conditions. We demonstrate how PTM-Shepherd improves the analysis of data from formalin-fixed paraffin-embedded samples, detects extreme underalkylation of cysteine in some datasets, discovers an artefactual modification introduced during peptide synthesis, and uncovers site-specific biases in sample preparation artifacts in a multi-center proteomics profiling study.

Deep proteome coverage in bottom-up proteomics requires peptide-level fractionation to simplify the complex peptide mixture before analysis by tandem mass spectrometry. By decreasing the number of co-eluting precursor peptide ions, fractionation effectively reduces the complexity of the sample leading to higher sample coverage and reduced bias towards high abundance precursors that are preferentially identified in data-dependent acquisition strategies. To achieve this goal, we report a bead-based off-line peptide fractionation method termed CIF or Carboxylate modified magnetic bead-based isopropanol gradient peptide fractionation. CIF is an extension of the SP3 (single-pot solid-phase-enhanced sample preparation) strategy and provides an effective but complementary approach to other commonly used fractionation methods including strong cation exchange (SCX) and reversed phase (RP)-based chromatography. We demonstrate that CIF is an effective offline separation strategy capable of increasing the depth of peptide analyte coverage both when used alone or as a second dimension of peptide fractionation in conjunction with high pH RP. These features make it ideally suited for a wide range of proteomic applications including the affinity purification of low abundance bait proteins.

Guo-qiang Bi
Dec 15, 1998; 18:10464-10472
Articles

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Prime Minister Narendra Modi will travel to Brazil to attend the annual G20 summit and to Nigeria and Guyana as part of a three-nation visit beginning November 16, the Ministry of External Affairs (MEA) said on Tuesday.

Prime Minister Narendra Modi will travel to Brazil to attend the annual G20 summit and to Nigeria and Guyana as part of a three-nation visit beginning November 16, the Ministry of External Affairs (MEA) said on Tuesday.

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hello, help me!

There are many change in the bump design. I want to design bump by APD.

The bump(die) is a stagger , create it by die generator. 

Because，the pin is not isometric. In order to RDL routing, so the bump is not isometric.

I move the symbol pin in APD symbol edit（as show in the picture）,  and selected symbol RBM write device file, write library symbol.

Export the bga text( bga text out) ,But the bump is not modified, the bump is still stagger.

Can you help me!

pitch2> pitch1

thanks

This is the 17th installment of The Rationalist, my column for the Times of India.

One of my favourite English words comes from chess. If it is your turn to move, but any move you make makes your position worse, you are in ‘Zugzwang’. Narendra Modi was in zugzwang after the Pulwama attacks a few days ago—as any Indian prime minister in his place would have been.

An Indian PM, after an attack for which Pakistan is held responsible, has only unsavoury choices in front of him. He is pulled in two opposite directions. One, strategy dictates that he must not escalate. Two, politics dictates that he must.

Let’s unpack that. First, consider the strategic imperatives. Ever since both India and Pakistan became nuclear powers, a conventional war has become next to impossible because of the threat of a nuclear war. If India escalates beyond a point, Pakistan might bring their nuclear weapons into play. Even a limited nuclear war could cause millions of casualties and devastate our economy. Thus, no matter what the provocation, India needs to calibrate its response so that the Pakistan doesn’t take it all the way.

It’s impossible to predict what actions Pakistan might view as sufficient provocation, so India has tended to play it safe. Don’t capture territory, don’t attack military assets, don’t kill civilians. In other words, surgical strikes on alleged terrorist camps is the most we can do.

Given that Pakistan knows that it is irrational for India to react, and our leaders tend to be rational, they can ‘bleed us with a thousand cuts’, as their doctrine states, with impunity. Both in 2001, when our parliament was attacked and the BJP’s Atal Bihari Vajpayee was PM, and in 2008, when Mumbai was attacked and the Congress’s Manmohan Singh was PM, our leaders considered all the options on the table—but were forced to do nothing.

But is doing nothing an option in an election year?

Leave strategy aside and turn to politics. India has been attacked. Forty soldiers have been killed, and the nation is traumatised and baying for blood. It is now politically impossible to not retaliate—especially for a PM who has criticized his predecessor for being weak, and portrayed himself as a 56-inch-chested man of action.

I have no doubt that Modi is a rational man, and knows the possible consequences of escalation. But he also knows the possible consequences of not escalating—he could dilute his brand and lose the elections. Thus, he is forced to act. And after he acts, his Pakistan counterpart will face the same domestic pressure to retaliate, and will have to attack back. And so on till my home in Versova is swallowed up by a nuclear crater, right?

Well, not exactly. There is a way to resolve this paradox. India and Pakistan can both escalate, not via military actions, but via optics.

Modi and Imran Khan, who you’d expect to feel like the loneliest men on earth right now, can find sweet company in each other. Their incentives are aligned. Neither man wants this to turn into a full-fledged war. Both men want to appear macho in front of their domestic constituencies. Both men are masters at building narratives, and have a pliant media that will help them.

Thus, India can carry out a surgical strike and claim it destroyed a camp, killed terrorists, and forced Pakistan to return a braveheart prisoner of war. Pakistan can say India merely destroyed two trees plus a rock, and claim the high moral ground by returning the prisoner after giving him good masala tea. A benign military equilibrium is maintained, and both men come out looking like strong leaders: a win-win game for the PMs that avoids a lose-lose game for their nations. They can give themselves a high-five in private when they meet next, and Imran can whisper to Modi, “You’re a good spinner, bro.”

There is one problem here, though: what if the optics don’t work?

If Modi feels that his public is too sceptical and he needs to do more, he might feel forced to resort to actual military escalation. The fog of politics might obscure the possible consequences. If the resultant Indian military action causes serious damage, Pakistan will have to respond in kind. In the chain of events that then begins, with body bags piling up, neither man may be able to back down. They could end up as prisoners of circumstance—and so could we.

***

Also check out:

Why Modi Must Learn to Play the Game of Chicken With Pakistan—Amit Varma
The Two Pakistans—Episode 79 of The Seen and the Unseen
India in the Nuclear Age—Episode 80 of The Seen and the Unseen

The India Uncut Blog © 2010 Amit Varma. All rights reserved.
Follow me on Twitter.

This is the 22nd installment of The Rationalist, my column for the Times of India.

Trade wars are on the rise, and it’s enough to get any nationalist all het up and excited. Earlier this week, Narendra Modi’s government announced that it would start imposing tariffs on 28 US products starting today. This is a response to similar treatment towards us from the US.

There is one thing I would invite you to consider: Trump and Modi are not engaged in a war with each other. Instead, they are waging war on their own people.

Let’s unpack that a bit. Part of the reason Trump came to power is that he provided simple and wrong answers for people’s problems. He responded to the growing jobs crisis in middle America with two explanations: one, foreigners are coming and taking your jobs; two, your jobs are being shipped overseas.

Both explanations are wrong but intuitive, and they worked for Trump. (He is stupid enough that he probably did not create these narratives for votes but actually believes them.) The first of those leads to the demonising of immigrants. The second leads to a demonising of trade. Trump has acted on his rhetoric after becoming president, and a modern US version of our old ‘Indira is India’ slogan might well be, “Trump is Tariff. Tariff is Trump.”

Contrary to the fulminations of the economically illiterate, all tariffs are bad, without exception. Let me illustrate this with an example. Say there is a fictional product called Brump. A local Brump costs Rs 100. Foreign manufacturers appear and offer better Brumps at a cheaper price, say Rs 90. Consumers shift to foreign Brumps.

Manufacturers of local Brumps get angry, and form an interest group. They lobby the government – or bribe it with campaign contributions – to impose a tariff on import of Brumps. The government puts a 20-rupee tariff. The foreign Brumps now cost Rs 110, and people start buying local Brumps again. This is a good thing, right? Local businesses have been helped, and local jobs have been saved.

But this is only the seen effect. The unseen effect of this tariff is that millions of Brump buyers would have saved Rs 10-per-Brump if there were no tariffs. This money would have gone out into the economy, been part of new demand, generated more jobs. Everyone would have been better off, and the overall standard of living would have been higher.

That brings to me to an essential truth about tariffs. Every tariff is a tax on your own people. And every intervention in markets amounts to a distribution of wealth from the people at large to specific interest groups. (In other words, from the poor to the rich.) The costs of this are dispersed and invisible – what is Rs 10 to any of us? – and the benefits are large and worth fighting for: Local manufacturers of Brumps can make crores extra. Much modern politics amounts to manufacturers of Brumps buying politicians to redistribute money from us to them.

There are second-order effects of protectionism as well. When the US imposes tariffs on other countries, those countries may respond by imposing tariffs back. Raw materials for many goods made locally are imported, and as these become expensive, so do those goods. That quintessential American product, the iPhone, uses parts from 43 countries. As local products rise in price because of expensive foreign parts, prices rise, demand goes down, jobs are lost, and everyone is worse off.

Trump keeps talking about how he wants to ‘win’ at trade, but trade is not a zero-sum game. The most misunderstood term in our times is probably ‘trade-deficit’. A country has a trade deficit when it imports more than what it exports, and Trump thinks of that as a bad thing. It is not. I run a trade deficit with my domestic help and my local grocery store. I buy more from them than they do from me. That is fine, because we all benefit. It is a win-win game.

Similarly, trade between countries is really trade between the people of both countries – and people trade with each other because they are both better off. To interfere in that process is to reduce the value created in their lives. It is immoral. To modify a slogan often identified with libertarians like me, ‘Tariffs are Theft.’

These trade wars, thus, carry a touch of the absurd. Any leader who imposes tariffs is imposing a tax on his own people. Just see the chain of events: Trump taxes the American people. In retaliation, Modi taxes the Indian people. Trump raises taxes. Modi raises taxes. Nationalists in both countries cheer. Interests groups in both countries laugh their way to the bank.

What kind of idiocy is this? How long will this lose-lose game continue?

The India Uncut Blog © 2010 Amit Varma. All rights reserved.
Follow me on Twitter.

hello:

i place a symbol into my design.

on my design, i change the symbol property by unlocking the symbol and unfixing pins so that i can move pins on the symbol.

i move some pins on my design.

but when i export the symbol from my design, the symbol is not current but has the original pin location.

is there a way to retain the pin locations after moving pins on a symbol when exporting the symbol?

regards

masa

Russia remains fDi’s most diversified commodity economy, while second ranked Brazil has displaced Ukraine into third place. Cathy Mullan reports.

At upcoming multilateral summits, Prime Minister Narendra Modi has an opportunity to expand India's regional position and economic links, and address issues such as terrorism and a rising China.

1. ‘Ek Hain Toh Safe Hain’: The Effect Of PM Modi's Slogan On Maharashtra’s Muslim Seats  News18
2. ‘Congress’s shahi parivar thinks it is born to rule India': PM Modi in Maharashtra  Hindustan Times
3. "Most Unstable Vehicle": PM Modi's Jibe At Maha Vikas Aghadi In Maharashtra  NDTV
4. Rahul Gandhi's 'Golden Goose' Is Revant Reddy | Claims of Congress Funding Accusation | India Today  India Today
5. Pune to progress faster in next five years, says PM Modi  The Times of India

to work with and modify the turbo hydra matic 40

to work with and modify the turbo hydra matic 400

α-Synuclein (α-Syn) aggregation in Lewy bodies and Lewy neurites has emerged as a key pathogenetic feature in Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy. Various factors, including posttranslational modifications (PTMs), can influence the propensity of α-Syn to misfold and aggregate. PTMs are biochemical modifications of a protein that occur during or after translation and are typically mediated by enzymes. PTMs modulate several characteristics of proteins including their structure, activity, localization, and stability. α-Syn undergoes various posttranslational modifications, including phosphorylation, ubiquitination, SUMOylation, acetylation, glycation, O-GlcNAcylation, nitration, oxidation, polyamination, arginylation, and truncation. Different PTMs of a protein can physically interact with one another or work together to influence a particular physiological or pathological feature in a process known as PTMs crosstalk. The development of detection techniques for the cooccurrence of PTMs in recent years has uncovered previously unappreciated mechanisms of their crosstalk. This has led to the emergence of evidence supporting an association between α-Syn PTMs crosstalk and synucleinopathies. In this review, we provide a comprehensive evaluation of α-Syn PTMs, their impact on misfolding and pathogenicity, the pharmacological means of targeting them, and their potential as biomarkers of disease. We also highlight the importance of the crosstalk between these PTMs in α-Syn function and aggregation. Insight into these PTMS and the complexities of their crosstalk can improve our understanding of the pathogenesis of synucleinopathies and identify novel targets of therapeutic potential.

Researchers have managed to encode enormous amounts of information, including images, into DNA at a rate hundreds of times faster than was previously possible

1. PM Narendra Modi to visit Nigeria, Brazil, Guyana from November 16  The Hindu
2. PM Modi to attend G20 Summit in Brazil, visit Nigeria and Guyana from November 16-20: Full schedule  The Times of India
3. PM Modi to embark on three nation visit to Nigeria, Brazil and Guyana from 16th nov  News On AIR
4. PM Modi's Africa-Latin America trip begins Sunday  The Economic Times
5. PM Modi to embark on 3-nation tour, attend G20 Leaders Summit in Brazil next week  The Indian Express