Cystic Fibrosis: New Hope for Gene Therapy?


Gene therapy to treat cystic fibrosis patients was associated with stabilization — but not improvement — in lung function, according to the results of a randomized, double-blinded phase IIb trial of the therapy in the U.K.

In a per protocol analysis, Professor Eric W.F.W. Alton, of Imperial College in London, and colleagues found a significant, but modest, treatment effect in cystic fibrosis patients treated with the nonviral, chemically designed gene-liposome pGM169/GL67A compared with those treated with a placebo of 0.9% saline (3.7%, 95% CI 0.1-7.3, P=0.046).

While a significant ANCOVA-adjusted effect was observed after 12 months’ follow-up, relative differences in FEV1 after 12 months of treatment were -0.4% (95% CI -2.8 to 2.1) for the treatment group compared to -4.0% (CI -6.6 to -1.4) for the placebo, they reported in The Lancet Respiratory Medicine.

The primary endpoint of the study was defined as relative percent change in forced expiratory volume (FEV1) after 12 months. The authors achieved that because while treatment with pGM169/GL67A did not improve lung function, it did not worsen it either.

Modest Benefit

“This study proves for the first time that copies of the normal CF gene delivered by aerosol inhalation can have a measurable beneficial effect on lung function, compared with placebo, in patients with cystic fibrosis,” senior co-author Dr. Alastair Innes, of Western General Hospital in Edinburgh, Scotland, told MedPage Today. He added that while the effect was statistically significant, he also described it as “modest.”

A small number of patients in both groups did experience improvements in lung function. The authors note that a post-hoc analysis showed 18% of patients (15 in the treatment group and six in the control group) showed an improvement in percent predicted FEV1 of 5% or more of their initial baseline values. By contrast, overall treatment effect in the 65 patients in the treatment group and 56 in the control group was 3.6% (95% CI 0.2-7.0,P=0.039). Of the 20 patients who did not complete the full treatment of one dose per 28 days for 12 months, they received a mean 3.7 doses (SD 1-9).

Patients were randomized into a number of stratified subgroups, but the authors attributed any treatment effect to a greater decline in FEV1 from the placebo group as opposed to greater improvement from pGM169/GL67A. Stratifying by baseline predicted FEV1 (<70% versus ≥70%) found that patients with a more severe disease (FEV1 49.6%-69.2% predicted) had a treatment effect of 6.4% (95% CI 0.8-12.1). By contrast, those with less severe disease (FEV1 69.6%-89.9% predicted) had a 0.2% treatment effect (CI -4.6 to 4.9, P interaction=0.065).

The authors also cited the post-trial and pre-trial changes in the placebo group (-4.9%) compared with the treatment group (1.5%) as contributing to the treatment effect. There were no differences observed by age, sex, or CTFR mutation.

The study also had a number of secondary outcomes, which achieved mixed results. Patients in the treatment group experienced greater improvements in forced vital capacity (FVC) and CT gas trapping, or the inability to exhale completely (P=0.031 andP=0.048, respectively) than the control group. But authors observed no treatment effect for other measures of lung function, imaging, and quality of life. Similar to the primary analysis, they did note that secondary outcomes tended to be more favorable for those with more severe disease.

The authors commented that patients with more severe disease seemed to experience an enhanced treatment effect, and saw this as an opportunity for further research.

“A larger trial with a stratified trial entry design, powered to assess subgroups, and that addresses the mechanisms of response heterogeneity will be important to verify or refute these data,” they wrote.

A total of six serious adverse events were recorded from the pGM169/GL67A group. The committee judged that they were unrelated to the treatment, though one may have been related to a trial procedure (bronchoscopy), the authors said. Two patients total discontinued treatment; one in the placebo group due to fatigue and one in the treatment group due to flu-like symptoms. There were no deaths during the study, and the authors saw no clinically relevant changes in patients throughout the study.

This randomized, double-blinded, placebo-controlled trial consisted of two cystic fibrosis centers in London and Edinburgh at 18 sites in the U.K. from June 12, 2012, to June 24, 2013. Participants were eligible if they were ages ≥12 years, had a FEV1 of 50%-90% predicted and had any combination of CFTR gene mutations. Of the 140 patients, 78 received pGM169/GL67A and 62 received a placebo. There were 116 patients (83%) completing the treatment and included in the per protocol analysis.

Limitations

The most important limitation the authors cite is that the mean difference is at the lower end of clinical trials for gene therapy in patients with cystic fibrosis, mainly due to the reduction in FEV1 volume in the placebo group. They suggest several reasons for this, such as optimal respiratory health for patients at time of trial entry, enthusiasm for the trial leading to improvements in lung function during the recruitment period, and that the trial included all available data, even if the patients were unstable, while registry data only contains measurements from an annual review. They also note the trial’s heterogeneous response and that the fact that changes may be the result of a “non-specific response” to the pGM169/GL67A treatment.

The authors describe their conclusions as a “proof of concept” for nonviral CFTR gene therapy, calling it “another step along the path of translational cystic fibrosis gene therapy.”

Innes said that the efficiency of the gene uptake needs to be improved before the therapy is applicable to clinical practice, adding that the UK Gene Therapy Consortium is engaged in pursuing several lines of research.

“We are exploring whether increased or more frequent dosing would increase benefit, the possible additional benefit of combining gene therapy with other basic treatments which help the CF ion channel to remain open, and novel viral gene therapy vectors which may increase the efficiency of gene transfer,” he said.

This Common Activity Is Reducing Your IQ, According to Research


Lack of sleep affects intelligence

The performance gap caused by an hour’s difference in sleep was bigger than the gap between a normal fourth-grader and a normal sixth-grader. Which is another way of saying that a slightly-sleepy sixth-grader will perform in class like a mere fourth-grader. “A loss of one hour of sleep is equivalent to [the loss of] two years of cognitive maturation and development,” Sadeh explained.
There is a correlation between grades and average amount of sleep.

Via NurtureShock:

Teens who received A’s averaged about fifteen more minutes sleep than the B students, who in turn averaged fifteen more minutes than the C’s, and so on. Wahlstrom’s data was an almost perfect replication of results from an earlier study of over 3,000 Rhode Island high schoolers by Brown’s Carskadon. Certainly, these are averages, but the consistency of the two studies stands out. Every fifteen minutes counts.
Not only does it affect intelligence, lack of sleep also reduces impulse control.

Via NurtureShock:

A different mechanism causes children to be inattentive in class. Sleep loss debilitates the body’s ability to extract glucose from the bloodstream. Without this stream of basic energy, one part of the brain suffers more than the rest—the prefrontal cortex, which is responsible for what’s called “Executive Function.” Among these executive functions are the orchestration of thoughts to fulfill a goal, prediction of outcomes, and perceiving consequences of actions. So tired people have difficulty with impulse control, and their abstract goals like studying take a back seat to more entertaining diversions. A tired brain perseverates—it gets stuck on a wrong answer and can’t come up with a more creative solution, repeatedly returning to the same answer it already knows is erroneous.
And when we’re tired it’s actually harder to be happy. We can recall negative memories more than positive ones when we’re exhausted.

5 HORRIBLE HABITS YOU NEED TO STOP RIGHT NOW
Do Not Email First Thing in the Morning or Last Thing at Night.“The former scrambles your priorities and all your plans for the day and the latter just gives you insomnia,” says Ferriss, who insists “email can wait until 10am” or after you check off at least one substantive to-do list item.
<strong>Do Not Agree to Meetings or Calls With No Clear Agenda or End Time</strong> “If the desired outcome is defined clearly… and there’s an agenda listing topics–questions to cover–no meeting or call should last more than 30 minutes,” claims Ferriss, so “request them in advance so you can ‘best prepare and make good use of our time together.’”

Do Not Email First Thing in the Morning or Last Thing at Night “The former scrambles your priorities and all your plans for the day and the latter just gives you insomnia,” says Ferriss, who insists “email can wait until 10am” or after you check off at least one substantive to-do list item.

Negative stimuli get processed by the amygdala; positive or neutral memories gets processed by the hippocampus. Sleep deprivation hits the hippocampus harder than the amygdala. The result is that sleep-deprived people fail to recall pleasant memories, yet recall gloomy memories just fine. In one experiment by Walker, sleep-deprived college students tried to memorize a list of words. They could remember 81% of the words with a negative connotation, like “cancer.” But they could remember only 31% of the words with a positive or neutral connotation, like “sunshine” or “basket.”

California legislators who passed SB 277 to force vaccines on children now want to do the same for adults with SB 792


Now that nearly all school-age children in California have been summoned into the state’s growing medical dictatorship through Senate Bill 277, corrupt politicians are now trying to pass a companion bill that would force adults to be vaccinated in accordance with the government’s prescribed vaccine schedule.

Introduced by California Senator Tony Mendoza of Senate District 32, Senate Bill 792 is the medical mafia’s latest attempt to force vaccines on individuals without informed consent. It would mandate that adults who work in settings where children are being cared for or educated to be jabbed with many or all of the same vaccines that the children are forced to receive, without the option for a personal or religious exemption.

Those who refuse the mandate would face criminal penalties for non-compliance, according to the bill’s verbiage, which takes it one step further than SB 277 in forcing people under duress to accept a medical treatment that they might otherwise refuse due to the risk of serious adverse events, including death.

“This bill would make California the first state to require mandated vaccinations for all childcare workers, including all private and public school early childhood education programs (Headstart, Private preK and preschools), family daycares, and daycare centers,” warns Vaccine Impact.

“This bill eliminates medical autonomy, crushes religious freedom, undermines personal freedom, and burdens quality providers with a non-optional series of medical interventions in the form of mandated vaccines that are not even 100 percent effective.”

SB 792 would force adults in childcare settings to get annual flu shots

If passed, SB 792 would require all qualifying adults to be vaccinated for influenza, pertussis, and measles beginning on September 1, 2016. These same individuals would also have to receive annual flu shots between August 1 and December 1 of each subsequent year, in perpetuity.

“This bill, commencing September 1, 2016, would prohibit a day care center or a family day care home from employing any person who has not been immunized in accordance with the schedule for routine adult immunizations, prescribed by the federal Centers for Disease Control and Prevention,” reads and excerpt from the bill.

“The bill would make conforming changes to provisions that set forth qualifications for day care center teachers and applicants for licensure as a family day care center. Because the bill would extend the application of a crime under the act, the bill would impose a state-mandated local program.”

Learn more: http://www.naturalnews.com/050595_SB_792_vaccine_mandates_medical_police_state.html#ixzz3hTHP96Ce

New Drug Lowers Levels of Triglyceride Blood Fats: Study


An experimental drug dramatically lowers blood levels of potentially harmful triglycerides, a new study finds.

Triglycerides are a type of blood fat created by the food you eat. At very high levels, they can cause heart problems and pancreatitis, an inflammation of the pancreas.

“Current treatment for elevated triglyceride [levels] leaves a lot to be desired,” said researcher Dr. Joseph Witztum, a professor of medicine at the University of California, San Diego. “This drug holds the promise that it will be the most effective therapy we have.”

The new drug — called ISIS 304801 for now — lowers triglyceride levels by as much as 71 percent without unpleasant side effects, the study found.

Elevated triglycerides can be caused by genetics as well as obesity, smoking, drinking too much alcohol and a diet very high in carbohydrates, the American Heart Association says.

Normal triglyceride levels are less than 150 milligrams per deciliter (mg/dL). But some people have much higher levels, which is associated with insulin resistance,metabolic syndrome, diabetes, inherited high cholesterol and certain other disorders, the researchers explained. Metabolic syndrome is a combination of high blood pressure, high cholesterol, high blood sugar and excess body fat.

A blood test will show levels of LDL (“bad”) cholesterol, along with HDL (“good”) cholesterol, and one-fifth of your triglyceride level, according to the American Heart Association.

Current treatments for elevated triglycerides include eating a healthy diet rich in fish oil and taking niacin (Niaspan), also called nicotinic acid. Drugs called fibrates, such as TriCor, can help lower triglyceride levels, but some patients don’t respond to the usual treatments, Witztum said.

The new drug works by targeting a protein — called apolipoprotein C-III — that slows the breakdown of triglycerides. In effect, ISIS 304801 speeds up the breakdown of triglycerides, allowing the fat to leave the body quickly.

“The drug lowers triglyceride and probably manifestations of insulin resistance and metabolic syndrome,” Witztum said.

For the study, researchers treated 57 patients with the drug or an inactive placebo. Their triglyceride levels ranged between 350 and 2,000 mg/dL, and they received weekly doses of the drug over 13 weeks.

The study also included 28 people who had triglyceride levels ranging from 225 to 2,000 mg/dL, who had been receiving fibrate therapy. These patients also received the new drug or placebo.

Overall, the researchers found that ISIS 304801 reduced triglyceride levels 31 percent to 71 percent.

This trial was the second of three required for drug approval in the United States. Witztum said that phase 3 trials are underway, but the results will not be available for two to three years.

Initially, the drug is intended to treat people who suffer from conditions such as inherited chylomicronemia syndrome that cause very high triglyceride levels, he said. In this condition, the body doesn’t break down fats correctly.

In the long run, Witztum believes the drug will be used by a larger population to help prevent heart disease related to high triglyceride levels.

The trial was funded by Isis Pharmaceuticals, the drug’s maker, and the results were published July 30 in the New England Journal of Medicine.

Dr. Gregg Fonarow, a professor of cardiology at the University of California, Los Angeles, said the study findings appear promising.

However, “whether lowering of triglyceride levels will in and of itself result in reduction in cardiovascular events remains to be demonstrated,” he said.

Further studies are needed, Fonarow said, to evaluate the drug’s safety and effectiveness, and determine whether it can prevent problems caused by high triglycerides.

“While the patient population that may benefit from a safe and effective triglyceride-lowering medication is more limited than the very broad population that have been shown to benefit from statin therapy to reduce cholesterol, there remains an important, but still unmet, clinical need,” Fonarow said.

5 Ways to Avoid Discomfort After Your Gallbladder Removal .


If you have your gall bladder removed, watch your diet carefully afterward. It can help you adjust gradually to changes in your digestion with little discomfort.

Gall bladder surgery solves the problem of gallstones, hard deposits of digestive fluid in the gallbladder. As people age, they become more common. In fact, surgeons remove more than 600,000 gallbladders each year to eliminate the pain associated with this condition.

The gallbladder – a pear-shaped organ on your right side beneath your liver – isn’t vital. However, it does help you digest fatty foods. It stores, concentrates and secretes the bile your liver makes.

After surgery, your liver still makes enough bile. But you might have difficulty processing fatty foods – at least for a while. More than half of patients who have their gallbladder removed have trouble digesting fat.

Here are five tips to avoiding discomfort after gall bladder removal:

1. Add foods back into your diet gradually

For the first few days after surgery, stick with clear liquids, broths and gelatin. After that, gradually add more solid foods back into your diet.

2. Go for low-fat and smaller portions

I advise my patients to avoid fried foods, high-fat foods, foods with strong odors and gas-causing foods. You should also stick to small, frequent meals.

Overall, I tell patients who have gallbladder removal surgery to stick to a low-fat diet. Typically, fat calories should total no more than 30 percent of your daily intake. That means if you eat about 1,800 calories each day, you should consume no more than 60 grams of fat.

Read food labels carefully. Look for foods that offer no more than 3 grams of fat per serving.

3. Skip high-fat foods t0 help avoid discomfort

Eating the wrong things after gallbladder surgery can induce pain, bloating and diarrhea. To side-step this gastrointestinal discomfort, avoid eating high-fat or spicy foods, including:

  • French fries and potato chips
  • High-fat meats, such as bologna, sausage and ground beef
  • High-fat dairy, such as cheese, ice cream and whole milk
  • Pizza
  • Lard/butter
  • Creamy soups and sauces
  • Meat gravies
  • Chocolate
  • Oils, such as coconut and palm oil
  • Chicken or turkey skin
  • Spicy foods

4. Take it slowly as you reintroduce high-fiber foods

Consider adding these gas-producing foods back into your diet slowly:

  • Whole-grain bread
  • Nuts
  • Legumes
  • Seeds
  • Brussels sprouts
  • Broccoli
  • Cauliflower
  • Cabbage
  • Cereal

Slowly add small amounts of foods back into your diet. Re-introducing things too quickly can lead to diarrhea, cramping and bloating.

5. Keep a journal and watch for ill effects

It’s a good idea to keep a food journal after surgery. You can keep track of when you begin eating a food again and what the impact is. Doing so will help you know what you can and cannot eat comfortably.

Most people can return to a regular diet within a month after surgery. However, talk to your doctor if you experience these symptoms:

  • Persistent, worsening or severe abdominal pain
  • Severe nausea or vomiting
  • Jaundice
  • No bowel movements for more than three days post-surgery
  • Inability to pass gas more than three days post-surgery
  • Diarrhea that lasts more than three days post-surgery

After surgery, doing these things should help you feel more comfortable. As time goes on, take note of your tolerance for higher fiber foods and fats, especially healthy fats.

Facebook Is About to Test Its Enormous Solar-Powered Drone


CX Deck

AT AN AIRFIELD somewhere in the UK, there’s a drone with the wingspan of a Boeing 737. And it belongs to Facebook.

This enormous unmanned aerial vehicle is called Aquila—a nod to the eagle who carried Zeus’s thunder bolts in Greek mythology—and it’s part of Facebook’s rather ambitious effort to deliver Internet access to the more than 4 billion people on earth who don’t already have it. The idea is that Aquila will circle in the stratosphere, above the weather, wirelessly beaming Internet signals to base stations in underdeveloped areas of countries like Nigeria and India.

Earlier this year, the company tested smaller models of this aircraft, and now, according to Facebook’s Yael Maguire, who oversees the project, the company is ready to test the full-size Aquila prototype. “The aircraft is real,” he tells WIRED, before a briefing with other reporters at Facebook headquarters in Menlo Park, California.

Though as wide as a 737, the drone weighs hundreds of times less than the commercial airliner, thanks to a carbon-fiber frame. The goal, Maguire says, is to reach a point where the drone can stay aloft for 90 days at an altitude of between 60,000 and 90,000 feet. “We think this is a very ambitious goal, given that the world record, as far as we can tell, is about two weeks.”

Meanwhile, at a lab in Woodland Hills, California, another group of Facebook engineers is developing new laser networking technologies that can help the drone beam its Internet signals down to earth. According to Maguire, the group has designed and tested a laser that can deliver data at “10s of Gbits per second,” hitting a target the size of a dime at a distance of 10 miles.

Developed under the aegis of a researcher group dubbed the Facebook Connectivity Lab, the Aquila project is just one of many efforts to deliver Internet access from the skies. Google is testing its own solar-powered drones—crashing one earlier this year—and it’s designing enormous balloons that can already stay aloft at similar heights for upwards of 180 days. Facebook and others are also exploring satellites that provide Internet signals from higher altitudes.

At Facebook and Google, these projects carry ulterior motives. The companies believe that if they expand the reach of the Internet, they’ll expand the reach of their businesses. Neither company plans to operate their own sky-high Internet services; they plan on handing these aerials to existing providers such as Vodafone. But these projects still have a long way to go. “There are many challenges ahead,” says Phil Finnegan, an analyst with the Virginia-based research outfit The Teal Group, who specializes in unmanned aerial vehicles. “Technologies are improving, but we’re not there yet.”

Indeed, it seems that Facebook has yet to test its 737-sized drone, and Maguire says the company likely won’t reach its 90-day-aloft goal until the end of this year or early next. At 60,000 feet, the drones are indeed above the most violent weather—and the clouds—but they must still deal with a little wind and extremely cold temperatures.

Though these planes are light and can power themselves via solar energy, Facebook must also find ways for the drones to carry all the equipment needed to deliver Internet signals, says Danny Ellis, the CEO of drone company SkySpecs, who has closely followed the project. Added weight could affect speed, but he believes that Facebook’s goal of a 90-day flight time is very doable. “Long-term, this is definitely a feasible idea,” he says. “We’ve already seen similar aircraft that can fly around the globe.”

Of course, Facebook must also find ways of reliably beaming that Internet access down to earth. Though the company is claiming that its laser technology is 10 times faster than the state of the art, Maguire declines to discuss the particulars of this technology. George Papen, an optical networking researcher at the University of California, San Diego, says it’s difficult to comment on Facebook’s claims because the company is not providing specifics, but he says that many others are working on similar technology, and that speeds that exceed 10s of Gigabits are well within what’s possible today. “This is not science fiction,” he says.

The real trick is to create a networking technology that can deal with obstacles such as clouds. “Guess what? There are clouds between there and Earth,” Papen says. In so many ways, this is a complex project. But Maguire and Facebook are very much committed to making it happen. The company’s social network now serves 1.3 billion people worldwide. But there are so many more to reach.

First Artificial Ribosome Designed


Scientists at the University of Illinois at Chicago andNorthwestern University say they have engineered a tethered ribosome that works nearly as well as the authentic organelle that produces all the proteins and enzymes within the cell. The engineered ribosome may enable the production of new drugs and next-generation biomaterials and lead to a better understanding of how ribosomes function, according to the researchers.

The artificial ribosome, called Ribo-T, was created in the laboratories of Alexander Mankin, Ph.D., director of the UIC College of Pharmacy’s Center for Biomolecular Sciences, and Northwestern’s Michael Jewett, Ph.D., assistant professor of chemical and biological engineering. The human-made ribosome may be able to be manipulated in the laboratory to do things natural ribosomes cannot do.

When the cell makes a protein, mRNA is copied from DNA. The ribosomes’ two subunits, one large and one small, unite on mRNA to form the functional unit that assembles the protein (translation). Once the protein molecule is complete, the ribosome subunits, both of which are themselves made up of RNA and protein, separate from each other.

In a new study (“Protein synthesis by ribosomes with tethered subunits”) in Nature, the researchers describe the design and properties of Ribo-T, a ribosome with subunits that will not separate. Ribo-T may be able to be tuned to produce unique and functional polymers for exploring ribosome functions or producing designer therapeutics, and perhaps one day even non-biological polymers, point out Dr. Mankin.” We felt like there was a very small chance Ribo-T could work, but we did not really know,” he noted.

Dr. Mankin said he and Dr. Jewett and their colleagues were frustrated in their investigations by the ribosomes’ subunits falling apart and coming together in every cycle of protein synthesis. Could the subunits be permanently linked together? The researchers devised a novel designer ribosome with tethered subunits, i.e., Ribo-T.

“What we were ultimately able to do was show that by creating an engineered ribosome where the ribosomal RNA is shared between the two subunits and linked by these small tethers, we could actually create a dual translation system,” explained Dr. Jewett. “It was surprising that our hybrid chimeric RNA could support assembly of a functional ribosome in the cell. It was also surprising that this tethered ribosome could support growth in the absence of wild-type ribosomes.”

Turns out that Ribo-T worked even better than Drs. Mankin and Jewett believed it could. Not only did Ribo-T make proteins in a test-tube, it was able to make enough protein in bacterial cells that lacked natural ribosomes to keep the bacteria alive. The researchers were surprised by this. Scientists had previously believed that the ability of the two ribosomal subunits to separate was required for protein synthesis.

“Obviously this assumption was incorrect,” said Dr. Jewett. “Our new protein-making factory holds promise to expand the genetic code in a unique and transformative way, providing exciting opportunities for synthetic biology and biomolecular engineering.”

“This is an exciting tool to explore ribosomal functions by experimenting with the most critical parts of the protein synthesis machine, which previously were ‘untouchable,'” added Dr. Mankin.

New Immunosuppressive Approved for Kidney Transplant Recipients


The FDA recently approved Veloxis Pharmaceuticals’ tacrolimus extended-release tablets (Envarus XR) to prevent organ rejection in kidney transplant patients. Veloxis anticipates that the product will be available in the United States in the fourth quarter of 2015. The FDA’s approval was based on the results of several late-stage trials that demonstrated once-daily Envarus XR has a flatter pharmacokinetic profile and higher bioavailability than twice-daily, immediate-release tacrolimus (Prograf), the current leading transplant drug.

“We are very pleased to have a new treatment option available for kidney transplant recipients,” said Anthony Langone, MD, associate professor and medical director of the Medical Specialties Clinic at Vanderbilt University, in a press release. “Patients must receive immunosuppression as lifelong therapy, and Envarsus XR holds promise for kidney transplant patients and their physicians as a tacrolimus product with the convenience of once-daily dosing.

” Envarsus XR and other immunosuppressants are associated with an increased risk for developing serious infections and malignancies that may lead to hospitalization or death. Other risks related to Envarsus XR include acute or chronic nephrotoxicity and mild to severe hyperkalemia. Hypertension is another common adverse effect of the drug and may require antihypertensive therapy.

Bacteria use DNA replication to time key decision


Bacteria use DNA replication to time key decision
This illustration of the replication cycle for circular bacterial DNA shows how Bacillus subtilis bacteria use the ratio of proteins KinA to Spo0F to time their decision to form spores. By copying the gene for Spo0F (purple) early in the cell-division cycle and the gene for KinA (green) later in the cycle, the bacteria assure that the decision to form a spore or divide is made when DNA replication has completed. Credit: L. Huang/Rice University

In spore-forming bacteria, chromosomal locations of genes can couple the DNA replication cycle to critical, once-in-a-lifetime decisions about whether to reproduce or form spores. The new finding by Rice University bioengineers and colleagues at the University of California at San Diego and the University of Houston appears this week in the journal Cell.

Like most microorganisms, Bacillus subtilis bacteria are single-celled creatures with one goal: to reproduce by making copies of themselves. But survival isn’t always that simple. For example, when food gets scarce, B. subtilismust decide between two possible paths: shut down, form a dormant spore—a process called “sporulation”—and wait for better times or split into two cells and gamble that there is enough food for at least one more generation.

“The decision about whether to form a spore and when is a very important one for B. subtilis,” said Oleg Igoshin, associate professor of bioengineering at Rice and one of the lead researchers on the new study. “If the organism waits too long, it can starve before it finishes transforming into a spore. If it acts too early and forms a spore too soon, it can be overwhelmed and out-reproduced by competitors.”

Igoshin’s lab specializes in describing the workings of the complex genetic regulatory networks that cells use to make such decisions. He said dozens of studies over the past 25 years have identified a network of more than 30 genes that B. subtilis uses to bring about sporulation. When food is plentiful, this network is largely silent. But during times of starvation the genes work in concert to form a spore.

B. subtilis is harmless to humans, but some dangerous bacteria like Bacillus anthracis, the organism that causes anthrax, also form spores by a similar mechanism. Scientists are keen to better understand the process, both to protect public health and to explore the evolution of complex genetic processes.

The exact workings of the sporulation network are complex. In 2012, Igoshin and graduate student Jatin Narula analyzed a genetic circuit downstream of the protein known as Spo0A, the “sporulation master regulator,” to explain how the network filters out noisy fluctuations in Spo0A activity. By filtering out noise, cells are able to accurately determine if Spo0A activity is above the threshold that triggers sporulation.

In the new study, Narula, Igoshin and collaborators set out to explain how B. subtilis times its sporulation decision with its cell-division cycle, a programmed series of events that cells normally follow to reproduce.

“Successful sporulation requires two complete copies of the bacterial chromosome, so coordination between the sporulation decision and the completion of DNA replication is very important,” Narula said. “A good analogy might be a semester-long course in biology. Lessons are presented in a particular order, and students are tested after they learn. If the final exam were given in the first week, students would almost certainly fail.”

Igoshin said that when the researchers set out to find how sporulation decisions were timed to the cell cycle, several studies including prior work by team members, provided a significant clue: Under starvation conditions, the activity of the master regulator gene had been shown to spike once per cell cycle.

In investigating how this spike occurred, Narula pored through dozens of published studies and noticed a discrepancy between some experimental results and the widely accepted view of the interactions between two key players in the sporulation network, a protein called Spo0F and a kinase called KinA. To resolve this discrepancy, Narula built a mathematical model in which excess Spo0F inhibits KinA activity. The new model showed that changes in the ratio of KinA to Spo0F could produce the pulse similar to those seen in experiments.

“The inhibition of KinA by 0F results in a ‘,’ which means the circuit output works to counteract the input that triggers it,” said Narula, co-lead author of the study. “Such loops are common in engineered and biological systems and usually work to keep things relatively constant despite external perturbations. A simple example of would be the thermostat on your house. When temperature drops it will keep your heater on until the temperature is back to normal. If there is a delay in the feedback loop, the system may overreact and produce a surge. With the thermostat, for example, if the heating unit continues to run for some time after the desired temperature is reached, the temperature can transiently spike before settling back to the desired level.”

Igoshin and Narula said similar spikes appear to be a consequence of the delayed negative feedback loop in the network that controls the amount of the active Spo0A. Furthermore, these spikes were timed based upon the positions of the KinA and Spo0F genes on the bacterial genome.

To divide and reproduce, bacteria must make a duplicate copy of their DNA. Because replication of circular bacterial DNA always initiates at one particular point, Narula surmised that the location of the KinA and Spo0F genes could be crucial. If one were located near the point where DNA replication began, the cell would contain two copies of that gene—doubling the rate of production of that protein—throughout the DNA replication period. If the other gene were located on the part of the circle that was copied last, the ratio of KinA to Spo0F would be one-to-one only when DNA replication was nearly completed.

Igoshin and Narula used a mathematical model of the network to show that this type of gene arrangement could account for spikes in Spo0A activity after each round of DNA replication. To verify their idea, they teamed with experimental biologists Anna Kuchina, co-lead author of the study, and Gürol Süel, co-lead investigator, both of the University of California at San Diego.

Experiments showed that the spikes of Spo0A activity always followed completion of DNA replication as the model predicted. In addition, Kuchina and colleagues used biotechnology to engineer mutant forms of B. subtilis in which the two critical genes were located near one another. The Spo0A spike from the delayed negative feedback loop was not observed in the mutants, and they failed to produce spores. In another engineered strain, the between Spo0A and Spo0F was eliminated. This led to a gradual increase in Spo0A activity as opposed to a spike, and such cells were several times more likely to fail or die during sporulation.

“We found that the relative location of sporulation genes on the DNA circle were similar in more than 30 species of spore-forming bacteria, including Bacillus anthracis,” Igoshin said. “This evidence suggests that the DNA timing mechanism is highly conserved, and it is possible that other time-critical functions related to the may be regulated in a similar way.”

NASA probe spots unusual red arcs on icy Saturn moon


Like graffiti sprayed by an unknown artist, NASA’s Cassini spacecraft has spotted unexplained arc-shaped, reddish streaks on the surface of Saturn’s icy moon Tethys.

The red arcs are narrow, curved lines on the moon’s surface, and are among the most unusual colour features on Saturn’s moons to be revealed by Cassini’s cameras, the US space agency said in a statement.

“The red arcs really popped out when we saw the new images. It is surprising how extensive these features are,” said Cassini participating scientist Paul Schenk from the Lunar and Planetary Institute in Houston.

A few of the red arcs can be seen faintly in observations made earlier in the Cassini mission.

But the colour images are the first to show large northern areas of Tethys under the illumination and viewing conditions necessary to see the arcs clearly.

The origin of the features and their reddish colour is a mystery to Cassini scientists.

Possibilities being studied include ideas that the reddish material is exposed ice with chemical impurities or the result of outgassing from inside Tethys.

They could also be associated with features like fractures that are below the resolution of the available images.

Except for a few small craters on Saturn’s moon Dione, reddish-tinted features are rare on other moons of Saturn.

Many reddish features do occur, however, on the geologically young surface of Jupiter’s moon Europa.

“The red arcs must be geologically young because they cut across older features like impact craters, but we don’t know their age in years,” added Paul Helfenstein, Cassini imaging scientist at Cornell University in New York.

The Cassini team is currently planning follow-up observations of the features at higher resolution later this year.

After 11 years in orbit, Cassini continues to make surprising discoveries, the statement read.