The Role of ADAR in Mammalian Development

Adenosine deaminase enzymes that act upon RNA (ADAR) have been shown to play a critical role in mammalian development.  ADAR is responsible for the conversion of adenosine to inosine in double-stranded RNA (dsRNA).

                    

Adenosine

Inosine

                                                          

Literally hundreds of thousands of these transforming events have been reported within human cells.  This editing appears to occur predominantly in noncoding repetitive elements within the genome.  Additionally, there are three forms of ADAR that have been identified – ADAR1, ADAR2 and ADAR3.  They seem to be widely expressed during early development especially in the embryonic and post-natal development period.  To date, of the three forms of ADAR, only ADAR1 and ADAR2 have been shown to demonstrate enzymatic activity on natural substrates in vitro.

Using the mouse animal model, ADAR2 has been shown to modify an essential receptor in the brain.  Experimental animals who die from seizures have successfully rescued when a genomic substitution mimicked the adenosine to inosine modification of the RNA transcript.  This result proved to be an elegant demonstration of the mode of action of ADAR2 in vivo.

The role of ADAR1 within the living organism remained unknown.  This prompted Dr. Brian J. Liddicoat and his colleagues from the Department of Medicine at St. Vincent’s Hospital University of Melbourne, Australia to engage in an elaborate experimental investigation to uncover its role.

The results of their extensive efforts demonstrated that, “The A-to-I editing of ADAR1 is essential for embryonic development and the maintenance of hematopoiesis in vivo.”  Hematopoiesis refers to the production of viable circulating blood cells.

This is an important finding and is a significant contribution to the overall understanding of the complexities and intricacies of mammalian development.

Climate Change Revisited

Despite the anti-science rhetoric that seems to have broad representation in the US Congress, climate change is a very real phenomenon that if unchecked will put the world's people in peril. If our collective behavior in regards to burning of fossil fuels continues unabated it will necessarily have a dramatic impart on peace and security in the world populated by future generations of human beings. The question we must pose to ourselves is, "Is this the legacy we want to leave behind?"


An example of what the future may have in store is what changes are occurring in the lives of the peoples of the Marshall Islands.

Somatic Mutation and Cancer

Cancer is a disease that can involve any tissue in the human body.  Some cancers, of course, are more common than others such as lung cancer, colon cancer, breast cancer in women and prostate cancer in men.  What all cancers share in common is that the tissue cell involved undergoes a transformation that imparts a selective advantage to that cell allowing it divide outside the normal controls imposed upon that particular cell type.  This process is known as clonal expansion.

It is now clearly understood that this selective advantage is a direct result of somatic mutation of the affected cell’s genetic material – DNA.  This causal relationship was first suspected in 1914 when it was observed that chromosomal abnormalities were present in cancer cells.  Note that this discovery was made even before DNA was known to be substance responsible for carrying hereditary information.    Considerably later, it was found that introducing DNA fragments from a malignant cell into a healthy one led to transformation of the recipient cell into a cancerous one.  Soon after this finding, oncogenes were discovered.  Oncogenes are the result of mutations in a class of genes responsible for cell growth or its modulation as is the case with tumor suppressor genes.

In nature, somatic mutations are common and are harmless for the most part – they do not generally impact genes that are involved in cellular process that would impart a selective advantage to the cell type affected.  These innocuous changes are referred to as “passenger mutations.”  Those rare mutations that do provide a selective advantage to the affected cell are referred to as “driver mutations.”  Ii is this type of mutation chat can lead to cancer – the production of a clone of cells that has the capacity to function autonomously and thereby no longer constrained by the controls normally imposed upon cells of a particular tissue.

Although this evolution of a cell from a normal state to one that is cancerous has been understood in general terms, the underlying process has not been amenable to discovery until the advent of high-throughput DNA sequencing. 

Utilizing this technology, the cancer-associated genes from over 10,000 human subjects have been sequenced along with over 2500 complete genomes from cancerous tissue samples.  Dr. Inigo Marincorena and associate from the Wellcome Trust Sanger Institute in Cambridge, UK have reviewed some of the result obtained from this data.

From this extensive study, new cancer genes have been discovered as well as new kinds of mutational events.    Mutations occur as a result of a number of possible factors.  These include:

• DNA replication errors
• DNA damage that is subsequently incorrectly repaired by the normal DNA repair machinery or damage that is left unrepaired
• DNA damage that is a result of exogenous factors such as mutagens, ultraviolet light and ionizing radiation
• DNA damage that is a result of endogenous factors such as reactive oxygen species (free radicals), aldehydes or mitotic errors
• Viruses and endogenous retrotransposons that lead to insertions into the genome.

It seems that the frequency of mutations found in various cancerous cell types vary depending upon the tissue involved.  For example, pediatric brain tumors and various types of leukemia have the lowest numbers of mutations whereas lung and skin cancers show the highest rates.  Furthermore these mutation rates can be greatly enhance if there is definitive loss of DNA repair pathways.

One of the conclusions drawn from the extensive genomic data derived from these studies is that more than one driver mutation is required in order for a clone originating from a transformed cell to successfully evolve into the fully cancerous state with the capability of metastasis.

This report represents a brief and limited overview of the extensive of this work and its significance.  It is fair to say, that future studies of this magnitude will shed greater light on the complex processes that are involved in cellular transformation and oncogenesis.

Climate Change – An Urgent Call to Action

Humanity does not seem to grasp the dim future that awaits the entire species if the dire warnings issued by climate scientists worldwide are not heeded regarding the absolute necessity to significantly diminish the burning of fossil fuels and thereby slow down the inexorable increase of the concentration of greenhouse gases in the atmosphere.

Ironically, human families work diligently towards fashioning a better life for their children, grandchildren and future descendants, yet their focus has been tilted towards material success and enhanced prosperity.  These are important goals.  However, if this onward rush towards greater and greater prosperity is not tempered by reality, the insidious consequences of climate change will make a significant portion of the planet essentially uninhabitable and rising seas will inundate the habitats of hundreds of millions of human populations wreaking havoc upon centers of economic power where great stores of human wealth reside.  Under such circumstances, such wealth would immediately be rendered quite meaningless.

It is quite absurd at this juncture to argue the merits of the science or allow short-term economic interests to resist the necessity for immediate and significant action.  If we do not act quickly as a world community, our descendants will find themselves marooned on what would increasingly become a hostile planet.  Is this the kind of future we are working so assiduously towards?

Obfuscation and resistance serves no useful purpose – the longer we procrastinate the more impossible it will be to make meaningful corrections on this our spaceship earth.

Ultimately, the science speaks for itself. 

Use of Genetically Modified Yeast in the Production of Opiates

Opiates are pain-relieving drugs that play a vital role in palliative care.  Morphine is a well-known member of this class of compounds.  Opiates have been derived from poppy plants for thousands of years.  The compound thebaine (paramorphine or codeine methyl enol ether) - see structure below -is the natural product that is the precursor for opiates.

Thebaine

Due to the incredible strides that have been made in regards to genetic engineering, a team of researchers headed by Christina Smolke from Stanford University in Palo Alto California has successfully incorporated genes derived from a variety of plants including poppies, bacterial genes from Pseudomonas and rodent genes into the yeast genome and endowed these modified yeast organisms with the capacity to produce thebaine from a sugar source.  This methodology required the successful incorporation of 21 different genes – a monumental enterprise.

This is a significant breakthrough in the area of synthetic biology for it suggests that eventually it will become possible to readily produce large quantities of thebaine that can subsequently be used to produce a variety of opiates.  Smolke and her colleagues avoided producing a modified yeast cell capable of producing morphine directly from sugar in order to forestall the possibility of inadvertently making the illicit production of morphine a practical reality.

The Role of Neutrophils in Guiding the Immune Response to Infection by the Influenza Virus

The influenza is virus has a global impact on human populations. Much research effort has been directed towards understanding how the human immune system combats influenza virus infection. The cellular target of this virus is the epithelial cells of the respiratory tract. In order for the immune system to successfully combat this pathogen, virus specific cytotoxic CD8+ circulating lymphocytes must migrate to the site of infection.

 It has been established that in order for these cytotoxic CD8+ lymphocytes to successfully kill virally-infected cells, it must be preceded by the innate immune response. A key player in this first response are Neutrophils (see image below) These cells are generally the first cell type to cross the blood vessel epithelium into the distressed tissue and are responsible for generating chemical signals that alert different types of immune cells.

 Although this relationship between Neutrophils and aspects of the adaptive immune response has been established, the underlying molecular mechanisms have been unclear. Dr. Kihong Lim and his colleagues at the Department of Microbiology and Immunology, David H. Smith Center for Vaccine Biology and Immunology, University of Rochester, Rochester, NY have investigated the mechanism of this immune response to infection by the influenza virus. What they uncovered was an interesting process.

Their studies have shown that the subset of cytotoxic CD8+ lymphocytes specific for influenza infected epithelium cells is dependent upon the presence of Neutrophils at the site of infection. The migrating Neutrophils leave in their wake a chemical trail; that chemical is a particular chemokine – a class of substances that serves as a powerful attractant to circulating lymphocytes – CXCL12. It is this CXCL12 that is a necessary requirement for the procurement of virus specific CD8+ lymphocytes at the infection site and for their subsequent ability to kill infected respiratory epithelial cells.

The results of this work contribute significantly to the fundamental understanding of the complex interactions that are required to mount a successful immunological assault upon a virus-induced infection.

Progress in Developing a Vaccine against the Ebola Virus

Progress in Developing a Vaccine against the Ebola Virus

The recent spread of hemorrhagic fever (EHF) in areas of West Africa including Guinea, Sierra Leone and Liberia has placed considerable urgency on the need to develop an effective vaccine against the pathogen responsible for this horrific and highly contagious disease, the Ebola virus (EBOV).  To date, of the approximately 27,200 reported cases, there have been more than 11,100 deaths – a mortality rate of  40.8 percent.   EBOV is so infectious that a high incidence rate has also been reported among health care workers.  It needs to be kept in mind that the health care infrastructure of the countries affected has been severely compromised especially since these so-called, “low resource” countries have budgets inadequate to respond effectively to this challenge.

By way of background EBOV belongs to a class of viruses called filoviruses.   Filoviruses are single-stranded RNA viruses.  This class of viruses possesses a glycoprotein (GP) on its surface that could conceivably make a good candidate as an immunogen – a protein capable of eliciting an immune response.

Dr. Andrea Marzi at the Laboratory of Virology, Division of Intramural Research at the National Institute of Allergy and Infectious Diseases  at the National Institutes of Health in Hamilton Montana.and his colleagues  have helped develop a strategy for developing such a vaccine.  The strategy they employed can be outlined in the following way –

• A live attenuated stomatitis virus was employed as a viral vector
• Recombinant technology was employed to modify this virus so that it expresses the Ebola GP on its surface referred to as Viral Stomatitis Vector EBOV (VSV-EBOV)
• This viral vector was then introduced into experimental animals – the rodent and macaque.

This approach was shown to be highly efficacious in both pre and post exposure vaccinations.  These results were so promising that phase 1 clinical trials in humans  were begun in several  worldwide locations. 

The following is in the author’s own words – “Complete and partial protection was achieved with a single dose given as late as 7 and 3 days before challenge, respectively.  This indicates that VSV-EBOV may protect humans against EBOV infections in West Africa with relatively short time to immunity, promoting its use for immediate public health responses.”