Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

Powerful New Technique to Reveal Protein Function

21.06.2013
In the cover story for the journal Genetics this month, neurobiologist Dan Chase and colleagues at the University of Massachusetts Amherst describe a new experimental technique they developed that will allow scientists to study the function of individual proteins in individual cell types in a living organism.

The advance should allow deeper insights into protein function, Chase says, “because we can only get a true understanding of what that single protein does when we isolate its function in a living organism. There was no tool currently available to do this.”

The journal’s cover art uses a jigsaw puzzle of a worm to illustrate how knockdown strategies in this organism have evolved over time to achieve more and more cell-type specificity, culminating in the new approach developed by the Chase lab, which can restrict knockdown to a single cell type.

“This strategy is super cool and it works great,” he says. “We’ve already used it to tease apart some of the mechanisms of dopamine signaling, but the strategy can be adapted to study the function of any protein involved in any biological process.”

There are more than 1 trillion cells in the human body, yet only 20,000 to 25,000 genes are expressed in them, Chase explains, so each gene must be expressed in many different cells. Understanding the function of 20,000 genes and whether this differs by cell type has been difficult, but over the last 10 years, he adds, “we have learned that the answer to this last question is a resounding yes. Gene function can differ by cell type.”

Pursuing this further, however, was hampered by the fact that traditional approaches for studying protein function rely on genetic mutations that act on DNA, so they disrupt protein function in ALL cells. And to understand what a protein really does, it must be studied in an individual cell in a living organism.

Specifically, Chase’s lab uses the roundworm C. elegans to explore how dopamine modulates the activities of specialized neurons. The worm is a useful model because it has only 302 neurons instead of billions in mammals. Despite its simplicity, the worm’s basic neurotransmission mechanisms are also found in humans.

In the quest to identify genes that regulate dopamine signaling, the UMass Amherst researchers quickly recognized that dopamine acts through proteins used by other neurotransmitters in other nervous system cells. “So we couldn’t use traditional genetic tools to study dopamine signaling. We needed to develop a new method to study protein function in individual cells in multicellular organisms,” Chase notes.

The technique they developed takes advantage of nonsense-mediated decay (NMD), a surveillance mechanism present in all eukaryotic organisms. NMD destroys aberrant mRNA molecules that can arise naturally through mutation during transcription or mRNA processing.

“In our strategy, we replace the normal copy of a gene with a tagged version that targets the gene’s mRNA for destruction by NMD,” Chase explains. “We then remove NMD from all the organism’s cells. Without NMD present, the replacement gene is expressed normally in all cells. We then knock down expression of the gene cell-specifically by restoring NMD activity only in cells we select.”

He adds, “This cell-specific restoration of NMD activity is easy and can also be controlled in time. Thus, using NMD we can not only remove gene function in individual cell types, we can control exactly when gene function is removed in that cell type. This gives complete control of gene expression and allows one to investigate the function of any gene in any cell type at any time.”

“With this very powerful new technique, now you can identify an individual gene and you can ask whether it plays a role in the behavior of interest. All these genes are expressed in our brains, so we are learning about all sorts of fascinating interactions in the worm and we can begin to translate the meanings to humans. Dopamine signaling is something you really can’t study in the human brain very well, but with this approach we are having success.”

To measure how much RNA is left in the animals after NMD is activated, microbiologist Aishwarya Swaminathan in John Lopes’ group at UMass Amherst used quantitative polymerase chain reaction (PCR), an amplification technique, allowing the researchers to precisely measure the efficiency of the strategy.

Chase says, “It turns out that the NMD-mediated knockdown is super good, better than anything else available. And anybody can use it, it’s straightforward molecular biology.”

Daniel Chase | Newswise
Further information:
http://www.umass.edu

More articles from Life Sciences:

nachricht Repairing damaged hearts with self-healing heart cells
22.08.2017 | National University Health System

nachricht Biochemical 'fingerprints' reveal diabetes progression
22.08.2017 | Umea University

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Fizzy soda water could be key to clean manufacture of flat wonder material: Graphene

Whether you call it effervescent, fizzy, or sparkling, carbonated water is making a comeback as a beverage. Aside from quenching thirst, researchers at the University of Illinois at Urbana-Champaign have discovered a new use for these "bubbly" concoctions that will have major impact on the manufacturer of the world's thinnest, flattest, and one most useful materials -- graphene.

As graphene's popularity grows as an advanced "wonder" material, the speed and quality at which it can be manufactured will be paramount. With that in mind,...

Im Focus: Exotic quantum states made from light: Physicists create optical “wells” for a super-photon

Physicists at the University of Bonn have managed to create optical hollows and more complex patterns into which the light of a Bose-Einstein condensate flows. The creation of such highly low-loss structures for light is a prerequisite for complex light circuits, such as for quantum information processing for a new generation of computers. The researchers are now presenting their results in the journal Nature Photonics.

Light particles (photons) occur as tiny, indivisible portions. Many thousands of these light portions can be merged to form a single super-photon if they are...

Im Focus: Circular RNA linked to brain function

For the first time, scientists have shown that circular RNA is linked to brain function. When a RNA molecule called Cdr1as was deleted from the genome of mice, the animals had problems filtering out unnecessary information – like patients suffering from neuropsychiatric disorders.

While hundreds of circular RNAs (circRNAs) are abundant in mammalian brains, one big question has remained unanswered: What are they actually good for? In the...

Im Focus: RAVAN CubeSat measures Earth's outgoing energy

An experimental small satellite has successfully collected and delivered data on a key measurement for predicting changes in Earth's climate.

The Radiometer Assessment using Vertically Aligned Nanotubes (RAVAN) CubeSat was launched into low-Earth orbit on Nov. 11, 2016, in order to test new...

Im Focus: Scientists shine new light on the “other high temperature superconductor”

A study led by scientists of the Max Planck Institute for the Structure and Dynamics of Matter (MPSD) at the Center for Free-Electron Laser Science in Hamburg presents evidence of the coexistence of superconductivity and “charge-density-waves” in compounds of the poorly-studied family of bismuthates. This observation opens up new perspectives for a deeper understanding of the phenomenon of high-temperature superconductivity, a topic which is at the core of condensed matter research since more than 30 years. The paper by Nicoletti et al has been published in the PNAS.

Since the beginning of the 20th century, superconductivity had been observed in some metals at temperatures only a few degrees above the absolute zero (minus...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

Call for Papers – ICNFT 2018, 5th International Conference on New Forming Technology

16.08.2017 | Event News

Sustainability is the business model of tomorrow

04.08.2017 | Event News

Clash of Realities 2017: Registration now open. International Conference at TH Köln

26.07.2017 | Event News

 
Latest News

Cholesterol-lowering drugs may fight infectious disease

22.08.2017 | Health and Medicine

Meter-sized single-crystal graphene growth becomes possible

22.08.2017 | Materials Sciences

Repairing damaged hearts with self-healing heart cells

22.08.2017 | Life Sciences

VideoLinks
B2B-VideoLinks
More VideoLinks >>>