mammalian cells

Molecule of the Day: Cisplatin

Cisplatin [Pt(NH3)2Cl2] is an metal complex that is used as an antineoplastic agent (anti-cancer drug). It is one of the archetypal transition metal complexes, being well-known and having a long history.

Cisplatin was first synthesised in 1845 by Michele Peyrone, and was known as Peyrone’s salt. Its medicinal effects, however, were not known until the late 20th century, until it was discovered by chance.

In 1961, a physics professor by the name of Barnett Rosenberg at Michigan State University embarked on a project to investigate the effect of electromagnetic radiation on cells during mitotic division. To his surprise, the setup caused the E. coli to elongate instead (below right), without undergoing cell division.

Eventually, the suspicion fell on the platinum electrodes. They were indeed the culprit; in the presence of oxygen, it had produced cisplatin. The researchers then decided to test its effects on mammalian cells by injecting it into mice with tumours; the results are remarkable, as can be seen from the images below. The tumour of the mouse that was injected with cisplatin (2nd row) had completely regressed by day 8, while the mouse that was not (1st row) eventually died on day 21.

Currently, cisplatin is used to treat various, but not all, forms of cancer, such as breast, lung, testicular, and ovarian cancer. When administered, the Cl atoms are displaced by water molecules, which themselves are displaced by guanine or adenine molecules in DNA. Since two ligands can be displaced, cross-linking of DNA occurs, interfering with cellular division.

Interestingly, its geometric isomer, transplatin, does not have any medicinal effect; it is believed to be due to its rapid hydrolysis when administered, causing it to be unable to react with the DNA to cross-link it. Additionally, the inter-strand cross-linking it causes could be more easily repaired by DNA repair mechanisms than the intra-strand cross-linking that cisplatin causes.

Cisplatin is synthesised from potassium tetrachloroplatinate via multiple ligand exchange reactions with potassium iodide, followed by ammonia, silver nitrate, and potassium chloride respectively.

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(this post is for @isaoubel​ as part of the [now closed] drive to help pwr bttm! they requested, “sufjan and aubrey meet solange.”

drake: solange, i can’t tell you how much i appreciate you taking the time to sit down and have dinner with us

solange: well, it’s always a pleasure to have… a seat at the table

sufjan: Oh Goodness Me She Said It Aubrey She Said The Name Of Her Record Did You Hear That Aubrey She Said It She Said A Seat At The Table Which Is The Name Of Her New Full Length Album A Seat At The Table She Said It She Said It She Said It

drake: yes sufjan she certainly did

solange: sufjan, if i may, i’d like to compliment you on your dinner party look. your jacket perfectly recalls the kineticism and vitality of keith haring and those wings… why, you appear positively apotheosized.

sufjan: I Am Going To Cry

drake: haha, don’t mind him… it just, it means a lot, coming from you. you’re sort of his fashion icon.

solange: that is so kind. thank you.

sufjan: I Love Your Dress It Makes You Look Like A Type Of Organelle In Eukaryotic Mammalian Liver And Gonad Cells That Forms An Interconnected Net Work Of Flattened Membrane Enclosed Sacs Or Tube Like Structures Known As Cisternae Which Functions In Lipid Manufacture And Metabolism As Well As The Production Of Steroid Hormones And Detoxification

solange: how perceptive of you; “smooth endoplasmic reticulum” is precisely the concept i aimed for.

sufjan: I Love You I Love Your Concepts And I Love The Synthesis Of Lipids

drake: i gotta be honest, i don’t understand what eighth-grade biology has to do with fashion

sufjan: With All Due Respect Aubrey You Frequently Perform In The Same Tee Shirts That You Wear As Pajamas… Your Performative Concepts Are Lacking

drake: my performative concepts? you’ve made some pretty ridiculous fashion statements yourself, sufjan! what about that time you headlined eaux claires in a trucker hat and pink walmart sunglasses and a gold chain?

solange: a trucker hat? pink walmart sunglasses? a gold chain? oh my!

drake: haha yeah! and he had this ridiculous tank top with, like, a tiger on it… and these big, like… cut-outs… along the sides… so everyone could see he’d been working out… and it was in the middle of july so he was all sweaty and-

sufjan: Aubrey Drake Stevens-Graham!

drake:

sufjan: 

drake: 

sufjan:

solange: ( ͡° ͜ʖ ͡°)

N6-methyladenine: A Newly Discovered Epigenetic Modification 

The majority of cellular functions are carried out by proteins encoded by specific genes present in cellular DNA. Genes are first transcribed to RNA which is then translated to proteins. The regulation of this process is important for maintaining correct cellular function. One of the ways that cells regulate gene expression is by epigenetic modifications to chromatin. The term “epigenetics” refers to reversible chemical modifications of DNA and histone proteins (DNA in the nucleus of eukaryotes is wrapped around histones) that affect the transcriptional status of genes. A number of histone modifications such as methylation and acetylation of lysine residues have already been discovered and characterized. Until recently; however, methylation of the 5 position of cytosine was the only known epigenetic DNA modification (A). Methylation of cytosine by DNA methyltransferases is associated with transcriptional silencing, while the removal of these methyl groups by TET enzymes is associated with transcriptional re-activation (B and C). In addition to controlling gene silencing, cytosine methylation also silences retrotransposons, a class of mobile genetic elements. If left unregulated, transposons can insert themselves into important regions of the genome and lead to mutagenesis.

Recently, N6-methyladenine, a new epigenetic modification, was discovered in mammalian cells. N6-mA had previously been discovered in prokaryotes and simple eukaryotes and was shown to function as a transcriptional activator. By contrast, a recent report published in Nature, has shown that N6-mA functions as a transcriptional silencer in mammalian cells, specifically in mouse embryonic stem cells. N6-mA primarily acts to silence the LINE-1 family of retrotransposons during early embryogenesis, which prevents genomic instability. The authors identified N6-mA by using a modified single molecule DNA sequencing technique. DNA bound to a specific modified histone protein was immunoprecipitated using an antibody against a specific histone modification (H2A.X), sequenced, and analyzed by mass spectrometry (D). This identified and determined the position of N6-mA. The authors then generated knockouts of the enzyme Alkbh1, which they believed may function as a demethylase for N6-mA. When Alkbh1 was absent from cells, they found an increase in the levels of N6-mA, showing that Alkbh1 functions as an N6-mA demethylase in vivo. This is important because epigenetic modifications are reversible. Genes can be turned off by methylation and then turned back on by removing the methyl group, so determining the enzyme responsible for the removal of N6-mA supports its role as an epigenetic modification.

For more information see:

http://www.nature.com/nature/journal/vaop/ncurrent/full/nature17640.html

As always, I’m happy to answer any questions or go into more detail.

Seeking SciNote, Biology: CRISPR

Question:

What do geneticists think will be possible when the the new gene-splicing CRISPR is fully operational on patients?

Answer:

For those of us unfamiliar, CRISPR is a revolutionary new genetic splicing technology. Gene splicing refers to modifications to a gene transcript that can result in different proteins being made from a single gene. Interestingly, CRISPR’s inception began when dairy scientists discovered that bacteria used to create yogurt (by transforming lactose into lactic acid) had incorporated snippets of benign viruses into its genome. To their surprise, the incorporated DNA would create toxic agents to thwart infective viruses. In 2007, dairy scientists realized that they could effectively fortify bacteria by adding spacer DNA, which does not code for any protein sequence, from a virus. Then, five years later, as Time Magazine writer Alice Park skilfully describes, professors Jennifer Doudna and Emanuelle Charpentier noticed “up to 40% of bacteria developed a particular genetic pattern in their genomes. What they found were sequences of genes immediately followed by the same sequence in reverse, known as palindromic sequences. Further, bits of random DNA bases cropped up after each such pairing and right before the next one. After the dairy bacteria transcribed its spacer DNA and palindromic sequence into RNA, it self-spliced those segments into shorter fragments, with an enzyme called CAS9”. As you may be wondering, CRISPR stands for “clustered regularly interspaced short palindromic repeats”.

It is important for us to emphasize the versatility of this method. In the 2007 article, Doudna and Charpentier go into depth regarding the many benefits of the new genetic technology. These include the potential to “systematically analyze gene functions in mammalian cells, study genomic rearrangements and the progression of cancers or other diseases, and potentially correct genetic mutations responsible for inherited disorders”. As you might imagine, this opens up possibilities that were previously science fiction. Currently, painful blood transfusions are commonplace in the treatment of many diseases such as sickle cell anemia. Sickle cell affects red blood cells, which are made by stem cells in bone marrow. Soon, Massachusetts Institute of Technology synthetic biologist Feng Zhang envisions that this will soon no longer be necessary. She predicts that after doctors extract some of the marrow, scientists will splice out the defective fragment of DNA using CRISPR from the removed stem cells, then bathe the cells in a solution containing the non-sickle-cell sequence. As the DNA repairs itself naturally, it picks up the correct sequence and incorporates it into the stem cell genomes. After this one-time procedure, the stem cells would give rise to more red blood cells with the healthy gene. Eventually, the blood system would be repopulated with normal cells.


The treatment of HIV using CRISPR would be very similar. In this potential treatment, “patients would provide a sample of blood stem cells from their bone marrow, which would be treated with CRISPR to remove the CCR5 gene, and these cells would be transplanted back to the patient. Since the bone marrow stem cells populate the entire blood and immune system, the patient would eventually have blood cells that were protected, or “immunized,” against HIV”.


Despite this extraordinary potential, no biological technology comes without serious ethical concerns. As Jennifer Douda says herself, CRISPR “really requires us to careful thought to how we employ such a tool: What are we trying to do with it, what are the appropriate applications, how can we use it safely?”

Check out her book The Stem Cell Hope for learning about the future of stem cell technology.

Sources:
Park, Alice. “A New Gene-Splicing Technique.” 100 New Scientific Discoveries: Fascinating, Unbelievable and Mind-expanding Stories. New York, NY: TIME, 2014. 92-95. Print.

Park, Alice. “It May Be Possible To Prevent HIV Even Without a Vaccine.” Time. Time, 6 Nov. 2014. Web.

Doudna, Jennifer A., and Charpentier, Emmanuelle (2014). The new frontier of genome engineering with CRISPR-Cas9. Science, 346(6213), 1258096–1258096. doi:10.1126/science.1258096

Answered by: Teodora S., Expert Leader and Expert John M.

Edited by: Carrie K.

nature.com
Light opens up the larynx
Muscles engineered to be photosensitive could lead to treatments for paralysis.

Scientists have genetically engineered muscles to move in response to pulses of light.

The technique, demonstrated on vocal cords removed from mice, is reported on 2 June in Nature Communications1. Researchers say that it could probe how muscles function — and might eventually help to treat people who have a paralysis that interferes with speech and breathing.

The work relies on a method called optogenetics, which can make cells that usually respond to electrical signals also react to light. The approach alters mammalian cells by inserting a gene for a protein such as channelrhodopsin, which in its natural setting allows blue-green algae to swim towards or away from light.

Optogenetics was first used in 2005 to modify neurons2, and has since become a standard tool to study the brain and nervous system. Applications outside neuroscience, however, are less common.

The latest study is fascinating, says Julio Vergara, a physiologist at the University of California, Los Angeles, who studies how electrical signals cause muscles to contract. “It shows the potential use of this very powerful technique for very important medical problems,” he says.

Continue Reading.