The gut-based tango of microbial and host genes

December 5, 2016 § Leave a comment

When you think ‘oscillations in your gut,’ you might think of motion sickness or food poisoning. But there is another type of oscillations in both the gut and other organs. These oscillations are an interplay of genetic switches for protein expression that turn on and off throughout the day as we transition through eating, working, exercising and sleeping. In a paper published in Cell on Dec. 1, researchers in Israel have investigated the links between the day-and-night circadian rhythm in mice and the microbes that thrive in their gastrointestinal tracts. They’ve found that the daily fluctuations that occur between the two systems are more intimately linked than previously expected.

The diurnal oscillations of microbial localization and metabolite production extend far beyond the gastrointestinal tract the microbes inhabit. Credit: Thaiss et al/Cell 2016

The diurnal oscillations of microbial localization and metabolite production extend far beyond the gastrointestinal tract the microbes inhabit.
Credit: Thaiss et al/Cell 2016

The systems close to one another but tightly coordinated, like a “tango between two partners,” says Eran Elinav at the Weizmann Institute of Science in Rehovot, Israel. Elinav and colleagues had previously linked shifts in our day-and-night circadian rhythm, such as jet lag, to disruptions of microbial gut communities in humans and mice that can lead to metabolic conditions, such as obesity and diabetes.

 

In their Cell paper, Elinav and colleagues, including Eran Segal from the same institute, homed in on the microbes that adhere to the epithelial cells of the gastrointestinal tract in mice with imaging and sequencing. They targeted the microbes’ entire genome. The targeting allowed the researchers to determine both the composition and function of the microbes.

They found that the bacteria residing in close proximity to the host lining epithelial cells displayed highly circadian behavior, with composition, function and microbial numbers differing at throughout a 24-hour cycle. Moreover, the thickness of the mucosal layer separating the gut bacteria from the mice’s epithelial layers fluctuated with the mice’s circadian rhythm and feeding patterns.

“Our findings also add to the increasing body of evidence that strongly suggests that disruption of proper circadian activity, such as that present in shift workers and frequent travelers, may drive metabolic derangements through a mechanism that is partly mediated by disruption of proper diurnal microbiome activity,” says Segal.

The investigators then wiped out these microbial communities with antibiotics to see how the mice’s transcriptome, the aggregate of genes being expressed via messenger RNA, adapted to the loss.

“There were a few hundred genes — the genes encoding the host clock itself — which did not care about the disruption to the microbiome,” says Elinav. “But there was another group of genes which normally oscillate in the host. Once we disrupted the gut microbes, these oscillations were completely lost.”

The investigators also noted that a subset of mouse genes that normally operate independently of the mice’s circadian oscillations began to follow the oscillations after the microbial communities were wiped out. These genes were picking up functions that had previously been performed by genes expressed by the microbiome.  “This brings the option that this “superorganism” shifts the tasks from one partner to the other once it is disrupted,” says Elinav.

The researchers were most shocked when they checked up on the mice’s livers. Despite the liver’s relative distance from the gastrointestinal tract, about 15 to 20 percent of its genes displayed circadian activity. “Surprisingly, when we disrupted the gut microbiome, the genetic program in the liver was severely disrupted,” says Elinav.

The researchers found that the metabolites, small molecules that are extensively modified by the microbiome and make up 80 percent of all the small molecules in peripheral blood, also displayed strong circadian activity. These molecules allow the gut microbiome to regulate the circadian activity in the liver.

When this was brought into the context of drug metabolism, the researchers found that liver toxicity induced by administration of high doses of the painkiller acetaminophen also displayed circadian activity. Interestingly, the researchers found that disrupting the gut microbiome reduced the toxicity of acetaminophen and stabilized it throughout a 24-hour period.

Elinav and colleagues are currently planning to continue investigating the intimacy of the gut microbiota and the effects of its diurnal activity in humans in order to elucidate potential systemic effects of antibiotics. They want to develop rational and safe intervention methods in the microbiome, potentially impacting human disease and drug metabolism.

This post was written by John Arnst, ASBMB Today’s science writer. 

Lifestyle chemistries of phones

November 22, 2016 § Leave a comment

Swabbing a phone for chemical signatures. Credit: Amina Bouslimani and Neha Garg, UCSD

Swabbing a phone for chemical signatures.
Credit: Amina Bouslimani and Neha Garg, UCSD

It used to be that the most troubling information you could get from swabbing someone’s phone case was an abundance of E. coli indicating his or her lack of good hygiene. In a paper published in Proceedings of the National Academies of Sciences on Nov. 14, researchers at the University of California, San Diego, expanded the scope of interrogation to include a number of trace chemical signatures. The signatures can give a picture of someone’s lifestyle.

“The number of molecules detected on every object will vary depending on the surface of the object and the lifestyle of these people,” says Amina Bouslimani at UCSD. Bouslimani is a postdoctoral researcher in the laboratory of Pieter Dorrestein and the first author on the PNAS study, which was funded by the National Institute of Justice, the research arm of the U. S. Department of Justice. “For every phone, we were able to detect between hundreds and thousands of molecules or compounds,” she continues.

Bouslimani and colleagues swabbed the phones and hands of 39 volunteers. They then paired mass spectrometry with a visualization process known as molecular networking. This allowed the researchers to group similar molecules and identify unknown molecules absent from a reference database based on their similarity to known compounds.

The researchers detected a 69 percent overlap between the samples taken from participants’ hands and the backs of their phones, which demonstrated a high transferability of chemicals between the two surfaces. Among many other food items, pharmaceuticals and hygiene products, the compounds detected corresponded to citrus fruits, caffeine, antidepressants, antifungal creams, hair-loss treatments, sunscreen and mosquito-repelling DEET.

The researchers also evaluated each participant’s potential exposure to flame-retardant plasticizing agents. They posited that this analysis could be used to monitor exposure to additional environmental hazards.

While the approach is not a replacement for DNA or fingerprint analyses, Bouslimani and colleagues hope that it might fill in gaps when DNA samples are contaminated or fingerprints recovered are only partials or not in a database.

“This work is exciting and very thought-provoking,” says Glen Jackson at West Virginia University, an expert in forensic analyses by mass spectrometry.

Jackson is cautious, however, about the accuracy of linking predicted activities with mass spectrometry-confirmed exposure to chemicals.

For example, while the presence of DEET based on data analysis may be very reliable information, he says, “proving that the lifestyle, or activity level, of the suspect is camping versus gardening is a different proposition altogether.” He added that there’s more work to be done to make sure that the results of such testing aren’t misconstrued.

The strength of the approach, according to Bouslimani, is the aggregate of the individual chemical signatures. “Our work flow doesn’t just detect one unique compound on this phone,” she says. “It is the combination of many such lifestyle chemistries that will help us to understand the personal habit and lifestyle.”

Bouslimani and colleagues hope to expand the breadth of their database, which would require the efforts of outside collaborators. “It has to be now a community effort,” she says. “We really hope that other people will start to apply this technology, to take this kind of development to the next level in forensic application.”

In the meantime, Bouslimani and colleagues plan to expand the study to include 80 people and each subjects’ keys, computers and wallets.

This post was written by John Arnst, ASBMB Today’s science writer. 

The most complete catalog of proteins in king cobra venom yet

June 30, 2016 § Leave a comment

King cobra at Kaeng Krachan National Park. Photo by Thai National Parks. Photo obtained from https://commons.wikimedia.org/wiki/File:Ophiophagus-hannah-king_cobra-kaeng-krachan-national-park.jpg

King cobra at Kaeng Krachan National Park. Photo by Thai National Parks. Photo obtained from https://commons.wikimedia.org/wiki/File:Ophiophagus-hannah-king_cobra-kaeng-krachan-national-park.jpg

Seven milliliters of a king cobra’s venom can kill 20 people. But what exactly is in the snake’s venom? Researchers have pursued that question for decades.

 

Now, in a paper published in the journal Molecular & Cellular Proteomics, a team of researchers reveals a detailed account of the proteins in the venom of king cobras. “I believe this study to be one of the most complete and precise catalogues of proteins in a venom yet obtained,” states Neil Kelleher at Northwestern University, one of the study’s senior investigators.

 

Snake venoms always have intrigued scientists, because they “have a rich diversity of biological activities,” says Kelleher’s collaborator Gilberto Domont at Universidade Federal do Rio de Janeiro in Brazil.  Among other things, venoms contain various proteases, lipases, nerve growth factors and enzyme inhibitors. Besides understanding how venoms function, researchers want to develop better antidotes to snake venom and identify molecules from venom that can be exploited as drugs, such as painkillers, anticlotting medications and blood pressure treatments. Domont points to captopril, a drug now commonly used to treat high blood pressure and heart failure. It was derived from a molecule found in the venom of a poisonous Brazilian viper.

 

Although the venom of the king cobra, the largest venomous snake in the world, which can stretch up to 13 feet, has been analyzed previously, questions persist about the venom. How do the sequences of the toxins evolutionarily vary? How do some post-translational modifications on proteins make the venom lethal? But to answer these questions, researchers need a proper count of the proteins in king cobra venom.

 

The advent of proteomics has allowed scientists to survey the rich diversity of proteins in a given sample. There are different approaches that rely on mass spectrometry to carry out proteomic analyses. One approach is called top-down proteomics. It allows researchers to look at proteins as whole, intact entities. In the more conventional approach, called bottom-up proteomics, proteins are cut into bite-sized fragments for analysis.

 

In bottom-up proteomics, researchers have to use computer algorithms to stitch back together protein fragments identified by mass spectrometry. Top-down proteomics avoids this problem. Its biggest advantage is that it can capture variations within the proteins as well as post-translational modifications.

 

Kelleher’s group is one of the leaders in developing top-down proteomics, so that’s what the investigators decided to use to analyze king cobra venom. Domont, Kelleher, Domont’s graduate student Rafael Melani and colleagues obtained venom from two Malaysian king cobras held at the Kentucky Reptile Zoo. They analyzed the venom by top-down proteomics in two modes, denatured and native. In the denatured mode, the protein complexes were taken apart; in the native mode, the venom was kept as is so the protein complexes remained intact.

 

The investigators identified 113 proteins in king cobra venom as well as their post-translational modifications. To date, only 17 proteins had been known in king cobra venom.

Why high-fat, low-carb diets help some epileptics

March 19, 2015 § Leave a comment

A series of pie charts depicting the calorific contributions from carbohydrate, protein and fat in four diets: the typical American diet; the Atkins diet during the induction phase; the classic ketogenic diet in a 4:1 ratio of fat to combined protein and carbohydrate (by weight); and the MCT oil ketogenic diet. Image from http://commons.wikimedia.org/wiki/File:Ketogenic_diets_pie_MCT.svg

A series of pie charts depicting the calorific contributions from carbohydrate, protein and fat in four diets: the typical American diet; the Atkins diet during the induction phase; the classic ketogenic diet in a 4:1 ratio of fat to combined protein and carbohydrate (by weight); and the MCT oil ketogenic diet. Image from http://commons.wikimedia.org/wiki/File:Ketogenic_diets_pie_MCT.svg

Antiepileptic drugs don’t work for one-third of patients. Instead, those patients usually eat ketogenic diets, high in fat and low in carbohydrates, which somehow stops seizures. In a paper just out in Science, researchers report how ketogenic diets work at the molecular level and say they designed a new drug that mimics the molecular effects of ketogenic diets.

“Current antiepileptic drugs are designed to target molecules that regulate electrical currents in neurons,” says Tsuyoshi Inoue at Okayama University in Japan, who led the work.  He adds that they “focused on antiepileptic actions of ketogenic diets” so they could find other ways to target epilepsy, particularly through metabolism.

Ketogenic diets force the body to rely on ketone bodies instead of glucose as an energy source. Inoue and colleagues explored what happens in single neurons in mouse brain slices when their energy source is switched from glucose to ketone bodies.

“We found that a metabolic pathway, known as astrocyte-neuron lactate shuttle, regulates electrical activities in neurons,” says Inoue.

An enzyme in the pathway is lactate dehydrogenase. When the energy source went from glucose to ketone bodies, the investigators realized that the switch in energy source inhibited the pathway via lactate dehydrogenase and caused the neurons to become hyperpolarized.

When the investigators inhibited lactate dehydrogenase in a mouse model of epilepsy, the animals suffered fewer seizures.

Next, the investigators used an enzymatic assay to see which existing antiepileptic drugs act on lactate dehydrogenase. They found a drug called stiripentol, used to treat a form of epilepsy called Dravet syndrome, inhibits lactate dehydrogenase. The investigators modified the drug’s chemical structure and found an analog that better suppresses seizures than the original.

Inoue says that inhibitors of lactate dehydrogenase can be “a new group of antiepileptic drugs to mimic ketogenic diets.”

Blog takes a break

August 13, 2013 § Leave a comment

Wild Types is taking a summer break! Check back at the end of the month for new posts.

Jon Lorsch to head National Institute of General Medical Sciences

March 25, 2013 § 1 Comment

Jon Lorsch, a professor at Johns Hopkins University, will be the next director of the National Institute of General Medical Sciences. He’ll arrive at the institute in Bethesda, Md., this summer.

Lorsch, an active member of the American Society for Biochemistry and Molecular Biology’s mentoring committee, will oversee a $2.4 billion budget that supports primarily fundamental research and scientific workforce training.

“With his reputation of being a broad-minded and visionary thinker with strong management skills, I am confident that Jon will lead NIH’s basic science flagship to keep the U.S. at the forefront of biomedical research,” Francis S. Collins, director of the National Institutes of Health, said in a statement announcing the appointment on March 25.

Lorsch will take the NIGMS reins from Judith H. Greenberg. Greenberg has served as the acting director of the institute since July 2011, when Jeremy Berg stepped down, after holding the director position for eight years, to become the University of Pittsburgh’s associate senior vice-chancellor of science strategy and planning.

Berg, who now is also president of the ASBMB, said he was pleased with the appointment: “Jon is a great choice. He is an outstanding scientist with ideas spanning many disciplines and with great teaching and training experience. He also led the curriculum reform efforts at Johns Hopkins and balanced clinical and basic perspectives very well.”

Berg continued: “He is very personable and is a good listener but is not at all afraid of tough issues. Jon is one of a small group of people whom I frequently reached out to when I was NIGMS director for his perspectives and advice.  NIGMS will be in good hands.”

Lorsch holds a Ph.D. in biochemistry from Harvard University and completed a postdoctoral stint at Stanford University. His group at Hopkins developed a fully reconstituted yeast translation initiation system, which the group used to understand the molecular mechanisms of the process by which the genetic blueprint in cells gets turned into working protein machines.

According to a Hopkins bio, Lorsch is thought to be the black sheep in his family “because, out of five males, he is the only one without a degree from Harvard Business School.”

ASBMB survey of young scientists sparks Twitter conversation about gender disparities

January 24, 2013 § Leave a comment

  1. Interesting note from bit.ly/UncG27 – Men outnumber women as applicants for faculty positions in biochem by almost 3:1. #WiS #STEM
  2. Survey suggests due in part to lack of knowledge of family-friendly policies… RT @Ilovepigenetics: @(me) Why are women leaving before TT?
  3. *********************************************************************************************************************
  4. @Ilovepigenetics @biochembelle Had a boss who’d let the women do the work, then take the credit, women never fought back ….
  5. @AjazSadia @Ilovepigenetics @biochembelle used to say if you were any good, you’d be raising family than working in the lab. So.
  6. @biochembelle That is the biggest sign about how TT is something I know has no interest to me based on how it’s set up. Despite the science
  7. *********************************************************************************************************************
  8. @biochembelle A TT job demands that your career comes first-TT women don’t have ‘wives’ at home making dinner. Whose career comes first?
  9. @dave_bridges @biochembelle I have seen many career focused women choose to follow their husband’s career. It’s more than dinner, housework.
  10. @Ilovepigenetics @dave_bridges Yep. Many surveys that say women still shoulder larger share of domestic duties in dual-career households.
  11. *********************************************************************************************************************
  12. .@biochembelle @Wikisteff We’ve got a majority of F in our biochem major here at AU. Fac used to be 1:1 M:F. After recent turnover is 4:2
  13. @biochembelle @dave_bridges Big disparity in # of female faculty. UofM A2 has many in powerful positions; other institutes not so much.
  14. @Wikisteff At our institution, Chemistry is 53% male over last three years. Institution is 43% male over same time. @biochembelle
  15. Though, I think there remain disparities in attitudes/pressures vs. official institutional policies. @biochembelle @Ilovepigenetics

Happy Holidays!

December 21, 2012 § Leave a comment

Image created by ASBMB's designer extraordinaire, Marnay Harris (http://marnayharris.com/)

Image created by ASBMB’s designer extraordinaire, Marnay Harris (http://marnayharris.com/)

The 12 days of Labmas

December 13, 2012 § 1 Comment

  1. SciTriGrrl
    @AmasianV @DrLabRatOry @biochembelle on a rhyming note: milkin’ = sulkin’? How many 9 postdocs sulkin’ ?@dr_leigh
  2. SciTriGrrl
    @biochembelle @DrLabRatOry @AmasianV the 5th day has to be ” five dead rats”! See how well that fits?#12daysoflabmas
  3. DrLabRatOry
    On the 3rd day of labmas my PI gave to me: 3 bench friends, 2 thermal gloves, and an old book of crystallography @AmasianV @biochembelle
  4. biochembelle
    On the 2nd day of labmas, my PI gave to me… 2 thermal gloves [both lefties] (and an old book of crystallography). @DrLabRatOry @AmasianV
  5. AmasianV
    @biochembelle on the first day of labmas my PI gave to me…

Urinary metabolic profile of clinical depression

November 19, 2012 § Leave a comment

Image courtesy of Kevin Dooley http://www.flickr.com/photos/pagedooley/2811155478/

Clinical depression, also known as major depressive disorder, robs its victims of interest and pleasure, sleep, appetite and concentration. Clinically depressed people also suffer from excessive fatigue and dark thoughts.  The illness is a major cause of disability, suicide and physical problems. However, a diagnosis for the illness is based on psychiatric reviews, which can be subjective. In a paper in Molecular & Cellular Proteomics, Chinese researchers described a test that could objectively diagnose the illness.

Depression is a complex mental disorder that involves multiple factors. The disease diagnosis is subjective because it can present a number of different symptoms and the exact causes for it are not understood. “Despite overwhelming efforts to identify the biomarkers for MDD, there were still no empirical laboratory tests available to diagnose MDD,” says Peng Xie of Chongqing Medical University who was the senior author on the MCP paper, adding that the current subjective diagnosis process has a considerable error rate.

The researchers decided to analyze urine, a sample that can be collected easily, for metabolites that could act as markers for depression. By using nuclear magnetic resonance spectroscopy, they were able to identify five molecules in urine that together seemed to sort out people who suffered from depression from those who didn’t.

The molecules were malonate, formate, N-methylnicotinamide, m-hydroxyphenylacetate and alanine. Malonate and formate are primarily involved in energy metabolism, m-hydroxyphenylacetate has a role in gut microbial metabolism and N-methylnicotinamide N-methylnicotinamide affects tryptophan-nicotinic acid metabolism. Alanine is one of the 20 amino acids used to make proteins. Xie says, “Based on the previous clinical and basic studies, we suggest that disturbances of these metabolic pathyways are implicated in the development of MDD.”

Xie says the researchers zoomed in on a few metabolites as markers because, in clinical practice, it is not convenient or economically feasible to simultaneously measure a large number of metabolites for diagnosis. The current work is a proof-of-concept and opens up more avenues of investigation. Xie says for one, the researchers would like to collect urine samples from depression patients and healthy controls from more ethnically diverse populations to further validate the diagnostic performance of the five metabolites. They also would like to dig deeper in to the underlying metabolic pathways of these five molecules to see if they can uncover how these biochemical pathways play into the disease.

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