Gene Mapping with Yeast

The human genome is often compared to a cell instruction manual – if each page were to represent one of the estimated 25,000 distinct genes, it would claim the title of the longest book ever to be written. These gene units are coded for by a unique fingerprint sequence of up to several million bases, each referred to by one of four basic letters: A, T, C or G. Despite this manuscript of life existing within most somatic cells, it was only in 2003 that the completion of the thirteen year-long Human Genome Project was able to cast light upon a genetic map that would later prove essential in navigating scientific research through the labyrinth of the genome. While the success of this project is widely lauded, the pivotal and unexpected role of S. cerevisiae (yeast) often remains in the shadows. 

The Importance of Yeast 

Despite the title of the Human Genome Project implying that research focused on human cells, yeast rapidly rose to the centre of mapping techniques for its unique properties that distinguished it from other candidates more similar to humans. While human genetic pedigrees – genetic trees used to display the Mendelian patterns of trait heritance - may be useful in analysis, the crevasse of time between generations stunted its potential applications and called for a faster-reproducing organism. The budding time of yeast averages 90 minutes, meaning that the trends in genetic composition and the occurrence of de novo (new) mutations from generation to generation could easily be observed. Furthermore, yeast has the potential to exist in both a diploid and a haploid form in relation to the environmental conditions; this permits researchers to initiate either sexual or asexual reproduction in a certain colony to monitor differences between these modes of replication. 

Tetrad Formation 

Yeast genes are mapped while it is in haploid – halved genetic material - form, requiring the yeast to sporulate under nitrogen-deficient conditions and create a tetrad of meiotically-divided haploids. This is performed via the following general method: 

The yeast sample is first streaked upon a petri dish and incubated, allowing for budding. Each colony appears as a distinct patch of yeast growth; a single colony is then isolated and swirled in a minimal media consisting of salts, minerals, a sugar source, and the absence of nitrogen. Under this lack of nitrogen, the yeast colony undergoes the evolutionary process of sporulation in response to stressful conditions to form an ascospore which would – in the wild – be able to drift to a more nitrogen-rich location. This is achieved as the cells exit the mitotic cell cycle of normal cell division and initiate meiosis within the nuclear envelope. During meiosis, the genetic material divides twice in succession, resulting in four daughter cells, known collectively as a tetrad. The membrane of the mother cell persists around the tetrad, acting as a protective ascus coating around the four inner spores.  

To reach the haploids for study, enzymes are employed for the dissolution of this ascus. The cells may then be observed using a powerful tetrad-dissecting microscope equipped with a fine glass needle designed to isolate the individual haploids from the tetrad.  

Using Tetrads to Measure Gene Linkage 

During the meiotic process, the genes do not segregate into identical cells as they would during mitosis. Instead, genetic variety of offspring is caused by recombination: this is the crossing over of DNA between different chromosomes to exchange genetic material at a certain point. Two genes are usually selected for observation to determine their genetic distance, and thus position within the yeast genome. The closer together the genes are, the more likely they are to remain on the same chromosome and in the same daughter cell following recombination.  

Without recombination, all haploids have what is known as the ‘parental ditype’ genotype; this is identical to that of the mother cell. A potential genotype of the mother cell could be AB/ab, in which A and a are two alleles of the same gene (as are B and b) and AB and ab represent the combinations of these alleles present on each chromosome belonging to a pair. If recombination does not occur between the loci of the two genes on the chromosomes, all haploid daughter cells have either an AB or ab genotype, which matches that of the mother chromosomes.  

However, in the event of recombination, one of two different offspring types may arise. The first is the non-parental ditype, in which none of the daughter cells have chromosomes that match the mother cell, as recombination has switched the arrangement of the two genes. This would be represented by a mix of Ab or aB haploids. The second is the tetratype, with four different possible genotypes – two of which are recombinant and two are parental. Therefore, the daughter cells would exhibit a mix of AB, ab, Ab and aB genotypes. Both these types of tetrads show that the chromosomes have crossed over and swapped material at some point between the genes A and B.  

Observing these haploids is critical in the measurement of genetic distance between yeast genes, since the relative numbers of each type of tetrad (parental ditype, non-parental ditype and tetratype) can be directly input into this formula, from which genetic distance measured in centimorgans (cM) can be derived:  

Genetic Distance = 100 x (T + 6NPD)/(2E) 

where T corresponds to the number of tetratypes, NPD to the number of non-parental ditypes and E to the total number of haploid cells in the sample. As recombination events increase in frequency, the numerator of the fraction rises since the number of tetratypes and non-parental ditypes increases in relation to the total number of cells. This causes the overall fraction to increase, displaying a proportional increase in genetic distance.  

Applications 

Although yeast seems an unlikely subject to map genetic distances and determine the degree to which genes are linked, it is to this unique organism that the Human Genome Project owes its success. While morphologically, humans and yeast are highly distinguishable, 23% of genes are homologous between these two species and observations of genetic distances in yeast are frequently mirrored in the human genome. These genetic distances can – like distances on a geographical map – be used to physically place the genes relative to each other to construct a highly accurate sequencing of bases.  

The most notable application of yeast technology resides in the study of genetic markers: these more visible and easily identifiable ‘flags’ are linked to and signal the presence of other, more significant alleles and mutations close by on the same chromosome. Among the marker loci identified using yeast are even genes which point towards antibiotic resistance in bacteria – a corner of research with the future potential to revolutionise healthcare and accelerate pharmaceutical evolution.  

References 

MITx 7.03.1 Genetics: The Fundamentals  

accessed: 19th November 2023 

https://www.uvm.edu/~dstratto/bcor101/mapping3.htm  

Accessed: 27th December 2023 

K-State Parasitology Laboratory: Mendelian Genetics Problems 

Accessed: 

https://www.k-state.edu/parasitology/biology198/answers2.html 27th December 2023 

A. Neiman: Ascospore Formation in the Yeast Saccharomyces cerevisiae 

Accessed: 

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1306807/#:~:text=The%20presence%20of%20a%20poor,%2C%20and%20sporulate%20(40). 27th December 2023 

A Glowing Review of 'Below the Edge of Darkness'

         Many months ago I found myself captivated by the idea of bioluminescence and spent many days researching it until every Wikipedia link had turned purple and I concluded my findings in a short article. After this short-lived love affair with light I returned to my non-luminescent world and this interest gradually became diluted by other discoveries and ideas I encountered in the following weeks. This was all to change after an animated afternoon discussion with a friend that led to me brushing the dust off my old notes about the deep ocean. At the reveal of my hidden interest, her eyes lit up as if they, too, were bioluminescent, and she insisted that I must at once read Widder's 'Below the Edge of Darkness'.

Never once have I read a non-fiction book disguised beautifully as a work of fiction. Widder is as much the protagonist of her piece as she is the writer. Typically as a reader of factual texts, I am left to feel as though I am sat hidden at the back of a standard, uninspiring lecture given by a professor with textbook-accurate fact recall. Widder instead seemed to reach a hand through the page and talk to me - as an older relative would an eager child - with the kind of wisdom only a life could give, not a textbook or a website. Just as it is perceived a sin to try to cleave art from artist, it is surprisingly hard to separate science from scientist. By the end of the book, I felt as though I knew Widder personally, and could see her human experiences shining through her discoveries. 

A particularly touching scene I found was the story of her close encounter with blindness. For a book clearly about light and the visible world, to open by plunging the reader into a shared period of both physical and emotional darkness was immensely impactful in developing an appreciation for the light we are so privileged to experience surrounding us. By prompting me to see even the everyday colour as beautiful and vibrant, she could then go on to truly dazzle with her descriptions of the unusual and breathtaking underwater scenes. And it was these depictions that were nothing like I had read before, even in fantasy novels. Widder takes us on a journey, bundling the readers into her claustrophobia-inducing submersibles and descending into the pelagic abyss. She skilfully manipulates our emotions to the point that - from the comfort of my bedroom - I felt my panic rise with hers as the precarity of these expeditions was unveiled and began to pray with her for a glimpse of the magical underwater world. At many points my heart momentarily halted as the world fell into an inky shadow. Then all of a sudden, the lights would flicker on and this array of colours would glow through the black and white pages like stars illuminating the night sky. I don't think I'll ever be able to look at the ocean in the same way again.

In terms of the science scaffolding these paintings, Widder has the perfect approach. Dealing with a subject you have decades of expertise in when communicating with the general public is rather like resurfacing after a deep-sea dive. You have to come up for air eventually and give an accurate and detailed narrative that represents your topic in an informative manner. But move too rapidly and abruptly and you risk decompression sickness. Not once did I find myself at a loss for detail while reading this. And not once did I find myself lost within the detail. It is clear that she not only has the wide knowledge about every facet of this topic, but also the intellect to express it in the best possible manner for her audience. And believe me, the audience was engaged. In fact, I had to check in the mirror after reading the final page that I hadn't begun to glow myself from the excitement for bioluminescence that Widder managed to rekindle within me!

The Legal Development of Genetic Testing

Recently, the development of genetic testing has given us greater control over our genes, from pre-implantation optimisation of in-vitro fertilisation to chorionic villus sampling. Some adult genetic tests even provide a window through which people can observe the future coiled within their own DNA, predicting changes that may lie dormant for as long as several decades. With this range of opportunity, however, comes a range of factors to consider including finance, ethics and legislation, and a deficit of information for those most concerned by it. 

        Perhaps the most widely-debated issue that genetic testing has created is the option of terminating gestation on the basis of results indicating a life-limiting disorder. Although this discussion comes with its own problems and ethical questions, a more overlooked topic surrounds adult genetic testing for later-onset conditions such as Huntington's disease. This is an autosomal dominant, neurodegenerative disorder resulting in chorea and mental decline between the ages of 30 and 50. The hidden implications of these genetic tests is explored by Bishop and Waldholz (1991, p322) in their book, Genome. These issues include the possibility of social and economic discrimination, such as potential biases against those with pre-diagnosed conditions in the workplace or by insurance companies maximising profit by exploiting those who will need the support most in the future. 

        Aside from the problems outlined by Bishop and Waldholz, other concerns also become apparent: what education and mental health guidance is on offer to those facing the results of these genetic tests? What is being done to ensure that people are able to make supported decisions regarding their results, and are fully informed during each section of the process? Since Genome was written over three decades ago, it acts as an interesting guideline to compare to today's legislatures and technology surrounding genetic testing. With this, we can view what has developed over recent years, and also what still remains to improve upon.

        In October 2018, the UK governmental Code on Genetic Testing and Insurance was put in place to tackle financial disadvantages experienced by many after receiving the results of genetic testing. The principle of the code is that insurance companies no longer have the right to require an applicant to undergo diagnostic or predictive genetic testing before receiving insurance, unless for a test relevant to the terms of the insurance itself. Furthermore, it is now against the law for insurance agencies to demand the results of a test of a blood-related family member, during the insurance cover, or when the result was obtained as part of a clinical trial. This is in the hopes that more people with a genetic history or predisposition to diseases such as Huntington's will feel more in control of their results and are able to make a better-informed decision to obtain a genetic test without the concern of data sharing and of bias based on their genetics. 

        The ability to recognise potential ethical and legal dangers in the field of testing has thus proven critical in its development: as much as genetic technology advances, so must our legislations.

Marine Bioluminescence

Since the first sparks of man-made fire ignited over 200,000 years ago, humans have been harnessing the power of light: from flames to filaments, this energy has guided us towards a world of innovation. However, it was only in recent eras of discovery that we learned that the true masters of light are those living deep underwater where the sun's rays cannot reach.  

How it Works

Marine bioluminescence is a phenomenon controlled on a molecular level, principally by an active (requiring adenosine triphosphate) chemical reaction in which oxygen combines with luciferin in a luciferase-catalysed process. Luciferins differ depending upon the species, with luciferases aiding the oxidation of a range of chemiluminescent substrates, emitting photons. Due to this photon release, a charged ion is a necessary element of the reaction as ionisation is often a requirement for electrons to shift to a higher energy level, thus emitting light energy as electrons return to their ground state.

On a Large Scale...

Bathocyroe fosteri - Marsh Youngbluth

Despite only being up to 40mm in length, the pictured resident of the mesopelagic zone is one of the larger bioluminescent aquatic invertebrates (the largest chemiluminescent marine animal being the kifetin shark at almost six feet.) On average, much of the light-emitting biomass is taken up by smaller plankton and bacterial colonies. Bathocyroe fosteri is a species of comb jelly utilising luciferins and luciferase as a combined photoprotein to emit shorter-wavelength visible light. The wavelength of this light and thus the colour is controlled by the size and the hydrophobicity of the attached isoleucine amino acid chain. Bathocyroe fosteri is located at depths of under 200m in the mesopelagic zone of the ocean but, despite their luminescence arguably improving hunting, this adaptation did not originally arise for this purpose. In the early atmosphere, oxygen is believed to have been highly toxic to organisms respiring anaerobically. Therefore, the oxidation of luciferin into the non-toxic product of oxyluciferin was an evolutionary adaptation to tackle this, with the side effect of chemiluminescence. It was long believed that comb jellies such as the Bathocyroe fosteri had no reception to light due to their lack of eyes. However, this luminescence may still prove useful since they possess photoreceptive opsins to detect light. 

...and a Cellular Level

Bioluminescent bacteria would serve little purpose emitting light on their own, only wasting valuable energy. Instead, they must light as a joined colony for the effect of the luminescence to be fully visible. This is achieved by quorum sensing, in which autoinducers become stimulated to trigger the oxidation of luciferins in the presence of a high concentration of bacteria in a specific region. Bioluminescent bacteria have become highly useful in the wider world of research, employing various purposes ranging from bioindication of aquatic pollutants to monitoring the distribution of genetically engineered bacterial populations released into ecosystems. It is also a possibility that in the future, these microscopic yet powerful species could even provide the key that unlocks solutions to sustainable urban light generation.

sources:

ocean ocean 6/28/23

science direct 6/28/23

bathocyroe 6/28/23

Neurogenesis: Could it Give us Clues in Battling Depression?

     Prior to the mid-20th century, adult neurogenesis - the generation of new nerve cells - was deemed to be impossible and unnecessary. Why would the developed brain require new neurons? Despite John Altman's significant discovery in 1965 of adult neurogenesis in rats, this question has remained without a concrete answer even to this day.

The process of neurogenesis is thought to occur in the hippocampus of the temporal lobe. This begins with the fabrication of dormant neural stem cells: slow-growing and multipotent cells which later divide into transit amplifying cells and transient intermediate progenitors (TACs and TIPs). These cells are much more rapidly-dividing and go on to create neuroblasts, cells which will soon differentiate into new neurons. This entire process lasts around 20 days and involves many activation hormones - the functions of which are still not entirely known - and stimuli specific to each individual type of neuron.

It is not just internal factors which promote neuronal growth; extrinsic stimuli have been proven to play a significant role in this process. These include: learning and working the memory, physical exercise, the surrounding environment and severe brain injuries. Lifestyle has an impact on the process of neurogenesis, but how does nerve growth affect our lives?

With the help of magnetic resonance imaging technology, a potential correlation has been witnessed between the instance of depression in patients and a decreased hippocampal size. This is a leading topic of discussion as it remains unclear whether the lack of neurogenesis is a risk factor for depression or if this reduction in capacity is just a symptom of mental decline. Studies carried out in rats point towards the latter, as while glucocorticoids (steroid hormones) have been proven to inhibit neurogenesis, when the inverse was tested with the blockage of neurogenesis, no change in mental stress was witnessed in the rodents. What this essentially means is that a lack of neurogenesis is simply a corollary of poor mental health and stress.

If depression leads to stunted regeneration, could stimulating neurogenesis have the opposite effect of preventing or even reversing mental health decline? Although this is not yet a theory which can definitively be proven, there is little harm in boosting physical activity and environment quality as a way to supplement well-being in patients and as a preventative measure in healthy people. Perhaps in the future the correlation will be more definitive and we may be able to improve the medications on offer to tackle the root cause of these conditions; for now neurogenesis remains an elusive - yet promising - topic of interest.

sources:

Science Direct 3/11/23

National Library of Medicine 3/11/23

Neuroscientifically Challenged 3/11/23

Behave, Robert Sapolsky 3/11/23