The Cat Genome is Finally Out of the Bag!

A Fishy Problem

For years, the genome of the friendly neighborhood Felis catus has been overlooked. After the human, mouse, and rat genomes were completed in the early 2000s, the NIH passed over cats in lieu of sequencing the dog genome. The question is, why? More influential people cared about dogs than cats. Furthermore, dogs suffer from many similar hereditary diseases as humans, such as arthritis. Kennel clubs also aided in the recruitment of many canine volunteers. Therefore, the canine genome was quickly sequenced. In fact, it was completed in 2005, two whole years before even an incomplete feline genome would be sequenced (1).

Cinnamon the Abyssinian cat was the first cat to ever have her genome sequenced. Image from the Cat Genome Sequencing Initiative.

The first cat to ever have her genome sequenced was an Abyssinian named Cinnamon. Light (2-fold) sequencing was completed in 2007. While inexpensive, this type of genome sequencing left many gaps. Approximately 60% of her DNA was sequenced (6). It wasn’t until 2014 that the first full feline genome was completed. Cinnamon’s genome was estimated to contain 19,493 genes that are thought to code for proteins and 1,855 noncoding RNAs (5).

Persian cats inherit PKD much more often than normal cats. Image by Flikr user Tavla.

Pawing for Feline Importance

Cats can serve as a valid models for many human diseases. Feline asthma, retinal atrophy, and type 2 diabetes are quite similar to their human counterparts. Cats suffer from several infectious diseases that are homologues to human diseases too, such as Feline Immunodeficincy Virus (FIV), the feline equivalent of HIV. Other diseases that cats suffer from that are also similar to human diseases are feline leukemia virus (corresponds to leukemia in humans), feline coronavirus (corresponds to avian influenza and SARS in humans), feline sarcoma virus (corresponds to sarcoma in humans). Therefore, cats are effective models of these diseases. (1, 6)

Cats also suffer from many hereditary diseases similar to those found in humans. For example, autosomal dominant polycystic kidney disease (PKD) is caused by mutations in the exact same gene in humans and cats (1, 3). PKD is the most prevalent hereditary disease in cats, since it affects approximately 6% of all cats in the world. It is found most predominantly in Persian cats, of which about 38% have inherited it. Afflicted cats suffer from cysts found in their livers, kidneys, and pancreases. The mutation that causes it is found in the PKD1 gene on the E3 chromosome. In this gene, Lyons et al. found a single transversion, in which a cysteine was replaced with an adenine. This single nucleotide change reads as a stop codon instead of another amino acid. Thus, translation of the protein the PKD1 gene codes for is stopped approximately 25% too early in cats with this mutation. It is thought that this single nucleotide polymorphism causes feline PKD. However, while this mutation has not been found in humans with PKD, similar areas of the resultant protein are missing. Consequently, cats are a viable natural model of this human disease. (3).

The Purrfect Solution

Fortunately, the Feline Genetics Lab at the University of Missouri is currently seeking to rectify this grievous imbalance. They have initiated a project called the 99 Lives Cat Genome Sequencing Initiative. Their goal is to sequence the genomes of 99 cats, both wild and domestic. Ultimately, this project will improve health care for cats and identify genetic variations within this adorable species. As of May 2015, they are over halfway there. Thus far, they have sequenced 53 individuals of varying breeds. (1, 4)

Lil Bub is a polydactyl cat, with more toes on each paw than normal cats. Image by Flikr user Kyle Matteson.

Lil Bub’s Big Genome

The next addition to this lineup is none other than the Internet sensation Lil Bub. She has several hereditary diseases. She has one extra toe on each paw, thus making her polydactyl cat. The scientists in charge of sequencing the Lilbubome have already discovered she is very distantly related to Ernest Hemingway’s famous polydactyl cats. She also may suffer from osteopetrosis, dwarfism, and mucopolysaccharidosis (either type I or IV), which are congenital bone disorders. Osteopetrosis is a very rare disorder that causes her bones to harden and grow denser. Mucopolysaccharidosis and/or dwarfism could be the cause(s) of her short stature. This project aims to identify the mutations that could result in her phenotype. (2)

References

1. Callaway, E. (2015). “I can haz genomes, too.” Nature, 517, 252-253. Retrieved May 11, 2015, from http://www.nature.com/news/i-can-haz-genomes-cats-claw-their-way-into-genetics-1.16708
2. Ibrahim, D., & Garcia-Lupiañez, D. (2015). The LilBubome – Sequencing LilBub’s Magical Genome. Retrieved May 11, 2015, from https://experiment.com/projects/the-lilbubome-sequencing-lilbub-s-magical-genome
3. Lyons, L. A., Biller, D. S., Erdman, C. A., Lipinski, M. J., Young, A. E., Roe, B. A., & … Grahn, R. A. (2004). Feline polycystic kidney disease mutation identified in PKD1. Journal Of The American Society Of Nephrology: JASN, 15(10), 2548-2555. Retrieved May 11, 2015 http://jasn.asnjournals.org/content/15/10/2548.full
4. Lyons, L. (n.d.). 99 Lives Cat Genome Sequencing Initiative. Retrieved May 11, 2015, from http://felinegenetics.missouri.edu/99lives
5. Montague, M. J., Li, G., Gandolfi, B., Khan, R., Aken, B. L., Searle, S. J., & … Warren, W. C. (2014). Comparative analysis of the domestic cat genome reveals genetic signatures underlying feline biology and domestication. Proceedings Of The National Academy Of Sciences Of The United States Of America, 111(48), 17230-17235. doi:10.1073/pnas.1410083111 Retrieved May 11, 2015 from http://www.researchgate.net/profile/Razib_Khan/publication/269094600_Comparative_analysis_of_the_domestic_cat_genome_reveals_genetic_signatures_underlying_feline_biology_and_domestication/links/54816dd80cf263ee1adfb8ce.pdf
6. Pontius, J. U., Mullikin, J. C., Smith, D. R., Lindblad-Toh, K., Gnerre, S., Clamp, M., & … O’Brien, S. J. (2007). Initial sequence and comparative analysis of the cat genome. Genome Research, 17(11), 1675-1689. Retrieved May 11, 2015 from http://genome.cshlp.org/content/17/11/1675.long

“Sorry, You Can’t Sit with Us, Your DNA is Mutated”- Genetic Discrimination and its Consequences

So, What Exactly is Genetic Discrimination?

http://www.clarksearch.com/

Genetic discrimination is the idea of using a person’s genetic information, and refusing to either insure, hire or treat someone based on that information. The majority of people concerned with genetic discrimination are ones that either have a genetic mutation or have family members who have a mutation. These mutations in a person’s DNA could lead to various genetic disorders, or potentially having susceptibility to other health risks. With having a higher amount of health risks, individuals fear that they may not get a job, or insurance, or even health care based on their susceptibility to these risks.

Genetic discrimination is a consequence of the push for genetic testing, which is a method to determine changes in a person’s chromosomes, genes or proteins. Through this testing, geneticists are able to determine the chance someone has in developing or passing on a certain condition or disorder. However, if someone’s genetic information is sequenced, there is a debate on whether or not other people should have access to that information, and if so, who gets to see it.

Repercussions of Genetic Discrimination:

Individuals are afraid that they can be denied health care based on their genetic information.
http://news-quality.com/

There are various different ways an individual can be genetically discriminated against. For the most part, people fear discrimination from health care insurance agencies, health care providers/ facilities, and possible places of employment. Let’s take me as an example, since I have a geneitic mutation that can lead to me being discriminated against. Having this mutation does not necessarily mean that I will develop certain health deffects that would prove to be expensive to cover, however, I do have a greater chance than someone who does not have a mutation. If an insurance agency or employer received my genetic testing showing my genetic mutation, I could be labeled as having a pre-existing condition. Because of this, I can denied either health insurance, which I need because of the nature of my disorder, or a job, if the employer does not want an employee that has a higher potential for a health risk. However, a majority of genetic mutations do not necessarily lead to a severe health deffect, and many people may not have a single complication in their life time.

So How Can We be Protected from this Discrimination?

http://www.geneticfairness.org/

Signed into law in 2008 by former President George Bush, the Genetic Information Nondiscrimination Act (GINA) prohibits discrimination based on an individual’s genetic information. Under Title I, the law prohibits the discrimination from health insurance companies. According to the law, it is illegal for any insurance company to force and individual to take a genetic test to determine the eligibility of being insured. Under title II of GINA, it is illegal to discriminate against a person based on their genetic information. Title II focuses on prohibiting discrimination from potential or current employers, and covers both current employees and any applicant of a certain company.

While this law does cover most instances of genetic discrimination opportunities, there is still some grey area that this law does not fully cover. For small companies (fewer than 15 employees), and certain military and veteran’s health insurance agencies, GINA does not apply. However, it is a start in the right direction to eliminate genetic discrimination.

This is Important, Why?

Discrimination against a person’s genetic information does little to advance our understanding of the human genome.For most mutations, there is no major health defect that occurs, however, there could be a slight risk of developing a defect. Individuals should not have to worry about not getting insurance or a job if they are considered to be more susceptible to certain defects. And just because someone may have a mutation, does not mean that they are any less of a human as the rest of society.

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Sources used:

http://ghr.nlm.nih.gov/handbook/testing/genetictesting

http://www.eeoc.gov/laws/statutes/gina.cfm

http://www.genome.gov/10002077

http://ghr.nlm.nih.gov/handbook/testing/discrimination

http://www.nlm.nih.gov/medlineplus/genetictesting.html#summary

http://www.geneticalliance.org/advocacy/policyissues/geneticdiscrimination

Jolie’s Overies Taken after she Fails Test

Well, not just any test, but arguably one more important than any math test you’ve taken. Ever. Since cancer is very prevalent in Jolie’s family (three female relatives died due to cancer), she decided to take preventative measures and have a test run to reveal her current chances of developing cancer. The first step was genetic testing, just as many other preventative surgeries begin. Her results showed a mutation in the gene attributed to a higher risk of breast cancer and ovarian cancer. After hearing this, there was no delay. Because of these genetic tests, and many others, people all over the world have been able to catch life-threatening conditions and live normal, healthy lives from then on out.

Testing Format

Today, there is about 900 genetic tests available for a multitude of different diseases, cancers, even certain phenotypes. The Genetics Home Reference defines three different types of genetic testing:

• “Gene tests”– tests that look at individual genes or DNA segments to find either slight variations or any mutations
• Chromosome tests-tests that focus only on chromosomes and whether or not there are any extras, if they are longer or shorter than they should be, or if any are missing
• Biochemical tests-use chemicals to measure protein activity where any changes could be an indication of a DNA change

Any one of these or a combination of them could end up uncovering any number of conditions or diseases. As for Jolie, the decision to remove her ovaries came after a test she took two years prior showed a mutation in the BRCA1 gene (this was when she had her mastectomy done…also in the news). Both BRCA1 and BRCA2 genes, when functioning normally, are responsible for making certain proteins that are know as “tumor supressors”. Essentially, they prevent a tumor from forming. But, if the gene is mutated such that the proteins do not work correctly or are not made at all, the chances of developing a tumor increase drastically.

How hard is this “test”?

Well, it depends on which test you are talking about! The testing for the BRCA1 and BRCA2 can be done just by a simple

So easy, your DNA can do it!

blood test. Other tests, like certain pre-natal screenings (Amniocentesis or Chronic villus sampling), pose a risk to the fetus and can only be done after a certain amount of gestation. In case you were wondering about if they have a test for a certain gene, phenotype or disease, The National Center for Biotechnology has a Genetic Testing Registry where you can search for tests based on what you actually want to test for. (Check it out…..it’s pretty amazing.) The search can tell you everything from how the test is done to who you can contact if you are considering the test.

Do you make the grade?

If you do, its a good chance its because of your parents. Although making the grade in this situation probably isn’t the best thing. In terms of the BRCA1 and BRCA2 genes, both of them can be inherited from your mother or your father. And, you chances of inheriting it are pretty high:

“Each child of a parent who carries a mutation in one of these genes has a 50 percent chance of inheriting the mutation.”

-National Cancer Institute

The National Cancer Institute states that by inheriting a mutated or damaged version of  the BRCA1 gene, females have a 55-65% chance of developing breast cancer by the age of 70. A bit lower if you inherited the mutated version of the BRCA2 gene-45%. The National Cancer Institute makes it clear that these percentages are not the same as they were, and may not be the same in some time; new information is always altering the percentages.

Do you want to know your results?

The big question. The open-ended essay question of genetic testing questions. So, I guess if you went and had a test done specifically for a certain gene or disease, you would want to know the results…that was the whole point of getting the test done. But what if you got a comprehensive gene test that looked at all of your DNA. Would you want to know if you had a gene for a life-ending condition? Or say that BRCA1 gene test cam back negative (meaning your genes are just fine) but something else showed up in the test…would you want to know then? Personally, I think I would have to side with finding out just what my genes have in store for me. Would you?

“Junk” DNA Is Not Actually Junk At All

What is “Junk” DNA?

Why Genomics

Junk DNA, a term first used in the 1960’s, was used to describe the non-coding sequences of DNA. According to News Medical, in a Nature review, Leslie Orgel and Francis Crick stated that junk DNA “had little specificity and conveys little or no selective advantage to the organism”. Scientists were unable assign any sort of function to these sequences, therefore, they thought this noncoding strand had no function.

According to PSRAST, if these sequences thought to be “Junk” DNA were in fact junk, the nucleotides in the DNA should be completely random, however, it has been found that the sequence of these are not random at all. Because of this, scientists now believe this “Junk” DNA must contain some kind of coded information.

What Does It Do?

Imgkid

Scientists now believe that this “Junk” DNA has various regulatory rules. This non-coding DNA influences the coding DNA in an important way. One reason scientists believe this DNA has regulatory functions is because the sequences of the “Junk” DNA are inherited and some repetitive patterns are associated with increased risk for cancer. This DNA can mutate rapidly in response to the cancer, so it has been speculated that it may contribute to the regulation of cellular processes (PSRAST).

According to PSRAST, there was a study done by Harvard Medical School on the “Junk” DNA. The study found that a “Junk” DNA strand regulated the activity of nearby genes. Most genes make proteins, but these genes work by being switched on. When the genes are switched on, it blocks the activity of an adjacent gene. A statement from the study was that “the researchers have evidence that the new gene, SRG1, works by physically blocking transcription of the adjacent gene, SER3.” Another study, called the ENCODE project, was conducted and the result was “three quarters of the non-coding DNA in the human genome did undergo transcription and that almost 50% of the genome was available to the proteins involved in genetic regulation such as transcription” (News Medical).

Cell Regulation

Riken Research

Many scientists believe that this “Junk” DNA helps regulate the cell and contributes to many human characteristics. When a gene is switched on, it can have a big impact on the size and shape of an organism. According to NPR, “there are many genes that have changed remarkably little between a mouse and a human, for example, and yet they behave differently within the cell, and that’s largely due to the way they are regulated differently.”

In another study from NPR, scientists were trying to find a gene sequence that involved brain development. When scientists believed they found such sequence they took the chimp sequence and the human sequence and injected it into mouse embryos. The study showed that the human DNA turned on gene activity in the neural stem cells. The mouse cells injected with the human sequence were approximately 12% bigger, before birth, according to a report in the journal Current Biology. What makes this study important was that this sequence that involves brain development was found on what was previously thought of as “Junk” DNA. This is just another study to support researchers idea’s that the “Junk” DNA has regulatory functions.

The “Junk” DNA may also play an important part in the creation of human limbs. A study was conducted at Yale where researchers looked for DNA sequences that were different in humans and other animals. According to NPR, scientists took this human DNA sequence that was different from other animals and injected it into a mouse egg. After injection, he wanted to see what cells became active as the embryo developed and he found that this sequence of “Junk” DNA looked like a regulatory gene because it seemed to switch genes on and off.

So, Is It Really Junk?

Many of these scientists agree that many of the differences between humans and other primates come from these “Junk” DNA sequences. Previously, these sequences were thought of as “Junk” because researchers could not find any function that these genes do. Now, with more technology and scientific advancements, scientists are now getting evidence that these sequences do, in fact, have some function, cell regulatory functions.

Synthetic Cells: The Future of Life

What is a synthetic cell?

Fir tree twig cross section at 10x magnification. Image by Flickr user Eckhard Völcker.

In many articles about synthetic cells, the words “artificial” and “synthetic” are often used interchangeably. However, in the scientific community it is necessary to distinguish between the two. Natural cells are those cells produced in nature, like human skin cells or the xylem and phloem cells of a plant. These cells are made without human intervention. Conversely, synthetic and artificial cells are made with human intervention. A synthetic cell is a cell with chemically synthesized pieces, such as DNA (4). An artificial cell is a cell that is completely man-made and mimics some of the functions of a cell, such as growth, communication, or signaling (5). Furthermore, a “living” artificial cell is defined as a cell that is fully synthetic and has all of the properties of a living cell (3). However, a “living” artificial cell has not yet been created.

 

The First Synthetic Cell

Mycoplasma mycoides. Image by Flickr user Minh Tuấn Nguyễn.

The first synthetic cell was made by scientists at the J. Craig Venter Institute in 2010 at a cost of $40 million (6). First, the researchers wrote out and edited the DNA of Mycoplasma mycoides, which consisted of over one million base pairs, as a computer file. The researchers even wrote their names into the DNA code, in order to track and claim ownership of these new cells. Then this file was sent to a company called Blue Heron Bio, which then synthesized hundreds of short pieces of DNA based on the templates sent to them. These pieces of DNA were then sent back to the researchers at the J. Craig Venter Institute, where they were assembled piece by piece within yeast cells. Once the finished genome was finally completely assembled, it was transplanted into a cell that had been emptied of most of its contents (4, 6). It is important to note that the cytoplasm and plasma membrane of this recipient cell were not synthesized. However, after the DNA was transplanted and the cells were allowed to replicate on a plate, the offspring did not have any of the original protein molecules that were in the initial recipient cell. The researchers found that the properties of the progeny cells were expressed just as though the whole cell had been synthesized, as if “…the DNA software [built] its own hardware” (4). This synthetic cell, the first of its kind, represented a breakthrough in the discipline of synthetic biology.

 

Designer Eukaryotic Chromosomes

Saccharomyces cerevisiae. Image by Flickr user AJC1.

Since 2010, there have been many advances in the creation of synthetic cells. In March of 2014, an international team of researchers successfully inserted a functional synthetic chromosome into a eukaryotic organism for the first time (7). This synthetic chromosome, named synIII, was based upon chromosome III of Saccharomyces cerevisiae (1). This chromosome is the smallest of this organism’s chromosomes and contains roughly 2.5% of S. cerevisiae’s genes (1, 7). Many changes to the genetic code of this chromosome were made. Several areas of the DNA were removed, including introns and tRNA genes. The resultant synthetic DNA molecule consisted of 272,871 base pairs, whereas the natural DNA molecule it was based upon consisted of 316,617 base pairs. The alterations did not cause any decline in fitness or replication time (1). The synthetic chromosome was then placed into a living yeast cell. This synthetic chromosome survived in the progeny of the cells through 125 generations (1, 7). This experiment represented great progress towards a completely artificial “living” cell (7).

So what are the implications of synthetic cells?

With such rapid advances in the creation of synthetic cells and the declining costs of DNA synthesis, researchers will soon be able to engineer new eukaryotic genomes with synthetic chromosomes that code for desired phenotypes and functions (1). These breakthroughs provide researchers with remarkable opportunities to study life and its significance. They have also given researchers fantastic opportunities to study and understand the genome of many different species. The day when completely artificial “living” cells are created is getting ever closer (2). The new era of synthetic biology has the potential to revolutionize life as we know it.

 

References

1 – Annaluru, Narayana et al. “Total Synthesis of a Functional Designer Eukaryotic Chromosome.” Science (New York, N.Y.) 344.6179 (2014): 55–58. PMC. Web. 18 Feb. 2015. <http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4033833/>.

2 – Bedau, Mark, et al. “Life After The Synthetic Cell.” Nature 465.7297 (2010): 422-424. MEDLINE with Full Text. Web. 18 Feb. 2015. <https://www.bu.edu/abl/files/nature_opinion.pdf>.

3 – Deamer, David. “A Giant Step Towards Artificial Life?.” Trends In Biotechnology 23.7 (2005): 336-338. MEDLINE with Full Text. Web. 18 Feb. 2015.

4 – Gibson, Daniel G., et al. “Creation Of A Bacterial Cell Controlled By A Chemically Synthesized Genome.” Science 5987 (2010): 52. Biography in Context. Web. 18 Feb. 2015. <http://www.sciencemag.org/content/329/5987/52>.

5 – Hammer, Daniel A., and Neha P. Kamat. “Review: Towards An Artificial Cell.” FEBS Letters 586.Seville special issue (2012): 2882-2890. ScienceDirect. Web. 22 Feb. 2015.

6 – Hotz, Robert Lee. “Scientists Create Synthetic Organism.” The Wall Street Journal. The Wall Street Journal, 21 May 2010. Web. 18 Feb. 2015. <http://www.wsj.com/articles/SB10001424052748703559004575256470152341984>.

7 – Vergano, Dan. “Scientists Move Closer to Inventing Artificial Life.” National Geographic. National Geographic Society, 27 Mar. 2014. Web. 18 Feb. 2015. <http://news.nationalgeographic.com/news/2014/03/140327-functional-designer-chromosome-synthetic-biology/>.

Flavr Savr Tomato

http://plantsinaction.science.uq.edu.au/

The genetically modified tomato went to U.S. market on May 21, 1994 known as the Flavr Savr. This transgenic tomato was no longer able to produce polygalacturonase(PG), which is an enzyme involved in fruit softening, due to an deactivated gene. Tomatoes are normally picked before ripening when they are still green and ripened artificially by ethylene treatment. The Flavr Savr tomatoes, however, are left to ripen on the vine and still have a long shelf life, which was thought to allow them to develop their full flavor.

The Gene

http://www.ebi.ac.uk

Scientists knew that polygalacturonase had the ability to dissolve cell wall pectin, which was the key to fruit softening. According to California Agriculture, “researchers at Calgene, Inc proposed to suppress PG accumulation in ripening tomatoes by introducing a reverse-orientation copy of the gene, an “antisense” copy designed to prevent or drastically reduce the formation of PG.” In 1987, Calgene researchers cloned a PG gene along with methods of transformation and regeneration. They inserted this PG antisense gene into the DNA of some tomatoes. The reason for inserting PG antisense gene was to reduce the amount of PG produced in the tomato. Data found that these tomatoes generated as little as 1% of the PG found in traditional tomatoes. The U.S. Food and Drug Administration approved the introduction of Kanamycin-resistance gene constructions needed to create the PG anitisense gene.

Is there a difference?

http://www.ebi.ac.uk

As with all genetically modified foods, there comes concerns. Calgene researchers tried to handle all concerns about the Flavr Savr tomato by doing studies. There were concerns about the Kanamycin resistance protein and allergic reactions, however, date showed that allergic reactions were highly unlikely. According to California Agriculture, data submitted by Calgene showed that the Flavr Savr tomato was indistinguishable in almost every aspect from the traditional tomato. There were only two ways in which two tomatoes were different. The first difference was that the fruit cell wall pectin degraded more slowly in the genetically modified tomato(this being the main point of making the new tomato). The second difference was that in the new tomato, the tomato paste had a higher viscosity. The only differences between the two tomatoes supposedly did not increase any risk, it just changed the taste.

The Disadvantages

http://www.nytimes.com

Although the demand for this tomato was high and remained high the entire time it was on the market, there was still many who opposed genetically modified foods. There is very little known about the long term effects of genetically modified foods, such as the Flavr Savr tomato. According to Actionbioscience, there have been acute toxicity studies conducted with male and female rats for the tomato. These studies claim that the tomato has absent toxic effects, but many people see flaws in the study. There was an unacceptably wide range of rat starting weights and no histology of the intestines was done. There were many factors that make the studies done invalid.

Where are they now?

http://www.monsanto.com

Calgene was very transparent with their processes and labeling. According to a video done by the New York Times, the company Monsanto bought out Calgene because they had patents to key technology and encouraged labeling. Monsanto denies these accusations, however, they are one of the leading companies in genetically modified foods who does not support labeling. Shortly after Monsanto bought out Calgene, the Flavr Savr tomato was shelved and has been ever since.