Nucleic acid sequence

Scorpion venom is a painkiller for the grasshopper mouse | Mo Costandi

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Researchers have identified the molecular mechanisms that make the grasshopper mouse resistant to scorpion venom.

Grasshopper mouse

A southern grasshopper mouse approaches and prepares to attack an Arizona bark scorpion. Photo: Matthew and Ashlee Rowe.

The bark scorpion is, according to Wikipedia, the most venomous scorpion in North America, wielding an intensely painful – and potentially lethal – sting that stuns and deters snakes, birds and other predators. People unfortunate enough to have experienced the sting say that it produces an immediate burning sensation, followed by prolonged throbbing pain that can last for hours.

But the grasshopper mouse is completely resistant to the bark scorpion’s venom. In fact, it actively preys upon scorpions and other poisonous creatures. As the film clip below shows, it responds to the bark scorpion’s sting by licking its paw for a second or two, before resuming its attack, then killing and eating the scorpion, starting with the stinger and the bulb containing the venom. Researchers have now established exactly why this is – paradoxically, the venom has an analgesic, or pain-killing, effect on the grasshopper mouse.

The animal’s secret lies in two proteins, the sodium channels Nav1.7 and Nav1.8, which are found in a subset of sensory nerve fibres called nociceptors. These cells express numerous other proteins that are sensitive to damaging chemicals, excessive mechanical pressure, and extremes in temperature, and have fibres that extend from just beneath the skin surface into the spinal cord.

The sensor proteins relay these signals to Nav1.7 and Nav1.8, which then change their structure in response, so that their pores, which span the nerve cell membrane, open up, allowing sodium ions to flood into the cell. This causes the nociceptors to generate nervous impulses, which are transmitted along the fibre into the spinal cord. From there, the signals are relayed to second-order sensory neurons, which then carry the signals up into the brain, where they are interpreted as pain.

Ashlee Rowe of the University of Texas in Austin and her colleagues started off by injecting scorpion venom, formaldehyde and salt water into the hind paws of southern grasshopper mice and common house mice, and compared their behavioural responses.

The house mice licked their paws furiously for several minutes after being injected with venom or formaldehyde, but not when they were injected with salt water. By contrast, the grasshopper mice seemed completely oblivious to the venom, and barely licked their paws at all after being injected with it. They found the formaldehyde to be far more irritating, and the venom actually reduced the amount of time they spent licking their paws when the two were injected together.

Next, the researchers isolated sensory neurons from both types of mice and grew them in Petri dishes. They then added scorpion venom to the dishes and used microelectrodes to measure the electrical activity of the cells. This showed that the venom strongly activated cells from the house mice, making them fire with rapid bursts of nervous impulses, but actually prevented cells from the grasshopper mice from firing. Further investigation revealed that the scorpion venom directly binds to, and potently inhibits, Nav1.8 sodium channels from the grasshopper mice, but not the house mice.

Rowe and her colleagues performed a final series of experiments to determine how this happens at the molecular level. They sequenced the Nav1.8 gene from the grasshopper mouse, and compared it to that of the common mouse, to identify multiple DNA sequence variations that confer insensitivity to scorpion venom. All the mutations encode amino acid residues in or around the pore region of the Nav1.8 protein, replacing neutral residues with acidic ones that are attracted to water.

As a result of these tiny structural changes, scorpion venom binds to Nav1.8 and switches it off, perhaps by plugging the pore or making it impermeable to sodium ions in some other way, thus blocking the transmission of pain signals into the spinal cord.

The researchers confirmed the importance of the pore region by using genetic engineering to replace this segment of the common mouse gene with the corresponding segment from the grasshopper mouse gene. This made the resulting protein resistant to the venom, whereas substituting the pore DNA sequence in the grasshopper gene with that from the common mouse gene rendered it highly sensitive to the venom.

The ability to detect pain is critical for survival, as it alerts organisms to potentially life-threatening injuries. Venomous creatures have capitalised on this, by evolving neurotoxins that inflict pain by activating nociceptors in one way or another, thus detering would-be predators from attacking again. The grasshopper inhabits the deserts of North America and Mexico, and probably evolved resistance to venom as a physiological adaptation, which enabled it to eek out an existence in such an extreme environment by feasting on venomous prey.

Previous work has identified Nav1.7 as a key player in pain signalling, and researchers have identified a number of rare mutations in the gene encoding it, which make people either completely or partially insensitive to pain. Drugs that block Nav1.7 activity could therefore be effective pain-killers, and various research groups have been researching and developing such drugs. The new findings identify Nav1.8 as another potential target, and provide another potential route for the development new analgesic drugs.

Secret Botulism Paper Published.

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The discovery of a new form of the deadly botulinum toxin gets published, but its sequence is kept under wraps until an antidote is developed.

In a publishing first, the sequence of a newly discovered protein is not divulged in papers announcing the finding. Researchers at the California Department of Public Health in Sacramento discovered the protein, a new type of the extremely dangerous botulinum toxin, lurking in the feces of a child who displayed the symptoms of botulism. They published their findings in two reports on the website of The Journal of Infectious Diseases, but absent from either paper was the DNA sequence of the protein, the eighth form of botulinum toxin recovered from the bacteriumClostridium botulinum. The move represents the first time that a DNA sequence has been omitted from such a paper. “Because no antitoxins as yet have been developed to counteract the novel C. Botulinum toxin,” wrote editors at The Journal of Infectious Diseases, “the authors had detailed consultations with representatives from numerous appropriate US government agencies.”

These agencies, which included the Centers for Disease Control and Prevention and the Department of Homeland Security, approved publication of the papers so long as the gene sequence that codes for the new protein was left out. According to New Scientist, the sequence will be published as soon as antibodies are identified that effectively combat the toxin, which appears to be part of a whole new branch on the protein’s family tree.

Researchers identify key proteins that help establish cell function

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Researchers at the University of California, San Diego School of Medicine have developed a new way to parse and understand how special proteins called “master regulators” read the genome, and consequently turn genes on and off.

Writing in the October 13, 2013 Advance Online Publication of Nature, the scientists say their approach could make it quicker and easier to identify specific gene associated with increased – an essential step toward developing future targeted treatments, preventions and cures for conditions ranging from diabetes to neurodegenerative disease.

“Given the emerging ability to sequence the genomes of individual patients, a major goal is to be able to interpret that DNA sequence with respect to disease risk. What diseases is a person genetically predisposed to?” said principal investigator Christopher Glass, MD, PhD, a professor in the departments of Medicine and Cellular and Molecular Medicine at UC San Diego.

“Mutations that occur in protein-coding regions of the genome are relatively straight forward, but most mutations associated with disease risk actually occur in regions of the genome that do not code for proteins,” said Glass. “A central challenge has been developing a strategy that assesses the potential functional impact of these non-coding mutations. This paper lays the foundation for doing so by examining how natural genetic variation alters the function of genomic regions controlling gene expression in a cell specific-manner.”

Cells use hundreds of different proteins called transcription factors to “read” the genome, employing those instructions to turn genes on and off. These factors tend to be bound close together on the genome, forming functional units called “enhancers.” Glass and colleagues hypothesized that while each cell has tens of thousands of enhancers consisting of myriad combinations of factors, most enhancers are established by just a handful of special transcription factors called “master regulators.” These master regulators play crucial, even disproportional, roles in defining each cell’s identity and function, such as whether it will be a muscle, skin or heart cell.

“Our main idea was that the binding of these master regulators is necessary for the co-binding of the other transcription factors that together enable enhancers to regulate the expression of nearby genes,” Glass said.

The scientists tested and validated their hypothesis by looking at the effects of approximately 4 million DNA sequence differences affecting master regulators in macrophage cells in two strains of mice. Macrophages are a type of immune response cell. They found that DNA sequence mutations deciphered by master regulators not only affected how they bound to the genome, but also impacted neighboring needed to make functional .

The findings have practical importance for scientists and doctors investigating the genetic underpinnings of disease, said Glass. “Without actual knowledge of where the master regulator binds, there is relatively little predictive value of the DNA sequence for non-coding variants. Our work shows that by collecting a focused set of data for the master regulators of a particular cell type, one can greatly reduce the ‘search space’ of the in a particular cell type that would be susceptible to the effects of mutations. This allows prioritization of mutations for subsequent analysis, which can lead to new discoveries and real-world benefits.”

Source:  University of California – San Diego

Scientist who mapped human genome says we will be able to ‘print’ alien life from Mars

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J. Craig Venter says the next revolution in genetics will come from synthetic biology, as we learn to design and ‘print’ organisms with computers.
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Scientists will soon be able to design and print simple organisms using biological 3D printers says J. Craig Venter, the scientist who led the private-sector’s mapping of the human genome.

Venter predicts that new methods of digital design and manufacture will provide the next revolution in genetic with synthetic cells and organism tailor-made to tackle humanity’s problems: a toolkit of sequenced genes will be used to create disease-resistant animals; higher yielding crops; and drugs that extend human life and boost our brain power.

These ideas have been outlined in Venter’s latest book ‘Life at the Speed of Light: From the Double Helix to the Dawn of Digital Life’, in which the geneticists asks the age-old question ‘what is life?’ before detailing the history – and future – of creating the stuff from scratch.

For Venter life can be reduced to “protein robots” and “DNA machines” but he also believes that technology will unlock far more exotic opportunities for creating life. The title of the publication refers to the idea that we may be able to transmit DNA sequences found on Mars back to Earth (at the speed of light) to be replicated at home by biological printers.

“I am confident that life once thrived on Mars and may well still exist there today,” writes Venter. “The day is not far off when we will be able to send a robotically controlled genome-sequencing unit in a probe to other planets to read the DNA sequence of any alien microbe life that may be there.”

Venter’s ideas may sound like science fiction but he has achieved comparable feats in the past. Frustrated by what he viewed as slow government-led efforts to sequence the human genome in the 90s, Venter raised private capital to create a rival effort under the company name of Celera

Fears that Venter and his backers would attempt to patent the genome spurred the US-led effort into action and global genes-race was sparked, with both sides eventually agreeing to announce their result one day apart in February 2001.

Venter parted ways with Celera in 2002 and founded the J.Craig Venter institute in 2006. In 2010 he and his colleagues at the institute announced that they had created the world’s first synthetic organism.

The team creating a bacterium genome from scratch and ‘watermarked’ it with custom DNA strings (these included an encoded email address) before transplanting it into another cell. The cell then began to reproduce, making it the first living species created by humanity.

Although such pioneering work frequently raises ethical questions over the danger of humanity ‘playing God’, Venter writes that he is not concerned with such concerns. In ‘Life at the Speed of Light’ he writes: “My greatest fear is not the abuse of technology but that we will not use it at all.”

Sequence-Based Discovery of Bradyrhizobium enterica in Cord Colitis Syndrome.

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BACKGROUND

Immunosuppression is associated with a variety of idiopathic clinical syndromes that may have infectious causes. It has been hypothesized that the cord colitis syndrome, a complication of umbilical-cord hematopoietic stem-cell transplantation, is infectious in origin.

METHODS

We performed shotgun DNA sequencing on four archived, paraffin-embedded endoscopic colon-biopsy specimens obtained from two patients with cord colitis. Computational subtraction of human and known microbial sequences and assembly of residual sequences into a bacterial draft genome were performed. We used polymerase-chain-reaction (PCR) assays and fluorescence in situ hybridization to determine whether the corresponding bacterium was present in additional patients and controls.

RESULTS

DNA sequencing of the biopsy specimens revealed more than 2.5 million sequencing reads that did not match known organisms. These sequences were computationally assembled into a 7.65-Mb draft genome showing a high degree of homology with genomes of bacteria in the bradyrhizobium genus. The corresponding newly discovered bacterium was provisionally named Bradyrhizobium enterica. PCR identified B. enterica nucleotide sequences in biopsy specimens from all three additional patients with cord colitis whose samples were tested, whereas B. enterica sequences were absent in samples obtained from healthy controls and patients with colon cancer or graft-versus-host disease.

CONCLUSIONS

We assembled a novel bacterial draft genome from the direct sequencing of tissue specimens from patients with cord colitis. Association of these sequences with cord colitis suggests that B. enterica may be an opportunistic human pathogen.

Souirce: NEJM

 

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Sequence-Based Discovery of Bradyrhizobium enterica in Cord Colitis Syndrome
BACKGROUND
Immunosuppression is associated with a variety of idiopathic clinical syndromes that may have infectious causes. It has been hypothesized that the cord colitis syndrome, a complication of umbilical-cord hematopoietic stem-cell transplantation, is infectious in origin.
METHODS
We performed shotgun DNA sequencing on four archived, paraffin-embedded endoscopic colon-biopsy specimens obtained from two patients with cord colitis. Computational subtraction of human and known microbial sequences and assembly of residual sequences into a bacterial draft genome were performed. We used polymerase-chain-reaction (PCR) assays and fluorescence in situ hybridization to determine whether the corresponding bacterium was present in additional patients and controls.
RESULTS
DNA sequencing of the biopsy specimens revealed more than 2.5 million sequencing reads that did not match known organisms. These sequences were computationally assembled into a 7.65-Mb draft genome showing a high degree of homology with genomes of bacteria in the bradyrhizobium genus. The corresponding newly discovered bacterium was provisionally named Bradyrhizobium enterica. PCR identified B. enterica nucleotide sequences in biopsy specimens from all three additional patients with cord colitis whose samples were tested, whereas B. enterica sequences were absent in samples obtained from healthy controls and patients with colon cancer or graft-versus-host disease.
CONCLUSIONS
We assembled a novel bacterial draft genome from the direct sequencing of tissue specimens from patients with cord colitis. Association of these sequences with cord colitis suggests that B. enterica may be an opportunistic human pathogen.
Source: NEJM

 

DNA: the ‘smartest’ molecule in existence?.

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DNA is the molecule that contains and passes on our genetic information. The publication of its structure on the 25th of April 1953 was vital to understanding how it achieves this task with such startling efficiency.

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In fact, it’s hard to think of another molecule that performs so many intelligent functions so effortlessly. So what is it that makes DNA so smart?

Multi-millennial survivor

For such a huge molecule, DNA is very stable so if it’s kept in cold, dry and dark conditions, it can last for a very, very long time. This is why we have been able to extract and analyse DNA taken from species that have been extinct for thousands of years.

Scientists have ‘resurrected’ blood protein from preserved mammoths after harvesting their DNA

It’s the double-stranded, double-helix structure of DNA that stops it falling apart.

DNA’s structure is a bit like a twisted ladder. The twisted ‘rails’ are made of sugar-phosphate, which give DNA its shape and protect the information carrying ‘rungs’ inside. Each sugar-phosphate unit is joined to the next by a tough covalent bond, which needs a lot of energy to break.

In between the ‘rails’, weaker hydrogen bonds link the two halves of the rungs together. Individually each hydrogen bond is weak – but there are thousands of hydrogen bonds within a single DNA molecule, so the combined effect is an extremely powerful stabilising force.

It’s this collective strength of DNA that has allowed biologists to study genes of ancient species like the woolly mammoth – extinct but preserved in the permafrost.

Our cells need to divide so we can grow and re-build, but every cell needs to have the instructions to know ‘how to be’ a cell.

Intelligent error correction

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The consequences of wrongly read or copied information can be disastrous and cause deformities in the proteins.

So as DNA replicates, enzymes carry out a proof-reading job and fix any rare errors.

They tend to repair about 99% of these types of errors, with further checks taking place later.

DNA provides those instructions – so a new copy of itself must be made before a cell divides.

It’s the super-smart structure that makes this easy. The ‘rungs’ of the DNA ladder are made from one of four nitrogen-based molecules, commonly known as A, T, G and C. These form complementary pairs – A always joins with T and G always joins with C.

So one side of the double-stranded DNA helix can be used as a template to produce a new side that perfectly complements it. A bit like making a new coat zip, but by using half of the old zip as a template.

The original side and the new one combine together to form a new DNA double helix, which is identical to the original.

Cleverly, human DNA can unzip and ‘replicate’ at hundreds of places along the structure at the same time – speeding up the process for a very long molecule.

Molecular contortionist

Two metres of DNA coils like a telephone cord to fit into each cell

DNA is one of the longest molecules in the natural world. You possess enough DNA, stretched out in a line, to reach from here to the sun and back more than 300 times.

Yet each cell nucleus must contain two metres of DNA, so it has to be very flexible. It coils – much like a telephone cord – into tight complex structures called chromatins without corrupting the vital information within.

DNA bases – vital rungs in the ladder

There are four different nucleotide bases in each DNA molecule:

  • Adenine (A)
  • Thymine (T)
  • Guanine (G)
  • Cytosine (C)

These small molecules join DNA together and encode our genetic information.

And despite being packed in so tightly, the genetic material can still be accessed to create new copies and proteins as required.

Human cells contain 23 pairs of chromosomes, with each containing one long DNA molecule as well as the proteins which package it. It’s no wonder DNA needs to be extremely supple.

Amazingly, this folded and packed form of DNA is approximately 10,000 times shorter than the linear DNA strand would be if it was pulled taut.

This is why we have the ‘luxury’ of having the plans for our entire body in nearly every cell.

Biological database

DNA storage

 

A research team has encoded data in artificially produced segments of DNA, including:

  • A 26-second snippet of Martin Luther King’s classic anti-racism address from 1963
  • A .pdf” of the seminal 1953 paper by Crick and Watson describing DNA’ structure

The total data package was equivalent to 760 kilobytes on a computer drive. Physically, the DNA carrying all that information is no bigger than a speck of dust.

Genes are made up of stretches of the DNA molecule which contain information about how to build proteins – the building blocks of life which make up everything about us.

Different sequences of the four types of DNA bases make ‘codes’ which can be translated into the components of proteins, called amino acids. These amino acids, in different combinations can produce at least 20,000 different proteins in the human body.

Think of it like Morse Code. It too uses only four symbols (dot, dash, short spaces and long spaces), but it’s possible to spell out entire encyclopaedias with that simple code.

Just one gram of DNA can hold about two petabytes of data – the equivalent of about three million CDs.

That’s pretty smart, especially when you compare it to other information-storing molecules. Using the same amount of space, DNA can store 140,000 times more data than iron (III) oxide molecules, which stores information on computer hard drives.

DNA may be tiny but with properties including stability, flexibility, replication and the ability to store vast amounts of data, there’s a reason why it must be one of the smartest known molecules.

With huge quantities of data being produced by ever-growing computer systems, traditional data storage solutions, like magnetic hard drives are becoming bulky and cumbersome. Researchers have now used DNA to store artificially-produced information, but could this be the future of data storage?

Source: BBC