DownHouseSoftware

My students know that I love horror. Especially horror films, in particular – and there are some films that I get obsessed with from time to time. My students know this because I let my love of horror bleed into my work and it can show up as decorations in my office, in the films I talk about before classes start, but most importantly in extra credit questions that I love to use to lighten the mood in exams and quizzes.

Tonight – Tuesday, September 27th, 2022, friends of mine and I will be watching and tweeting about a classic film, The Horror of Dracula (1958, available on HBOMax and Youtube) starting at 7pm Central. This is a great film, with a stellar cast including the incomparable duo of Christopher Lee and Peter Cushing. A must for any fan of horror or film in general.

Share tweets using the hashtag: #FullMoonFrights

The Fall semester is getting rolling again with a new class of Biotech first-years starting my molecular class and the seniors moving into their capstone work as well as my classes on Ethical Research and Bioinformatics.

This is the time when I collect errata to include as extra credit from Supreme Court Justices to the scientists who developed our basic notions of cell theory and the central dogma to my list of essential horror films.

In an effort to make these questions into a fairer opportunity to score extra points, I’m thinking of ways to balance my sources of questions. In the past, I’ve offered an array of questions across all of these genres (with an equal or greater opportunity to earn points coming from biology questions), but all questions were available to students. This year, I’m considering offering a choice of questions from which students can select from a list of say, for example, matched biology and horror questions such as the following:

Extra Credit: For each number, answer either question A or B. Circle the letter for the answer that you want to be graded (1 pt. each).

There are definitely more films that I want to add here, but it’s already a longer list than I’d like.

Scientists every biology student should be aware of (Molecular Biology Edition):

1. Gregor Mendel
2. Louis Pasteur
3. Walter Sutton and Theodor Boveri
4. Thomas Hunt Morgan (and Alfred Sturtevant)
5. Reginald Punnett
6. William Bateson
7. Charles Darwin and Alfred Russel Wallace
8. Eduard Buchner
9. George Beadle and Edward Tatum
10. Alfred Hershey and Martha Chase
11. Theodosius Dobzhansky
12. Matthew Meselson and Franklin Stahl
13. Erwin Chargaff
14. Frederick Griffith
15. Oswald Avery
16. James Watson, Francis Crick, Rosalind Franklin
17. Linus Pauling

Again, hardly an exhaustive list, but these are some who have received special mention in my Molecular class over the years.

I hope these lists and this new extra credit format work for my students and allow them to always be learning- even when it is just answering the question, “do you like scary movies?“

Over the years, I have commonly found that about 10-20% of my students have difficulty isolating DNA from their buccal (Cheek) samples. This typically occurs when centrifugation of these isolates (even at very low speed) intended to pellet cells prior to lysis and DNA isolation, results in the immediate lysis of cells and release of DNA into the saline solution. Sometimes this goes without the student noticing or appreciating what is happening in the tube and leads to a failure to obtain any detectable (by UV) or amplifiable (by PCR) DNA in their final tube.

Just today, Dr. Jerry Coyne (Faculty at the University of Chicago, author of ‘Why Evolution is True‘ and host of a blog of the same name), reported that the buccal sample he sent to 23andMe had a similar outcome.

They write to him, “Our laboratory tried to extract DNA from your sample, but unfortunately the concentration wasn’t high enough to meet our standards. While it is uncommon, it does happen occasionally due to biological variability between people.”

I still don’t know why some individuals’ cells appear fragile in this method, but perhaps it’s not entirely unique.

It’s time for final exams again and this year’s extra credit theme has been the classic Alfred Hitchcock product-…

Well, let’s just let the man himself tell us.

Astute students have undoubtedly watched this film already, but in case they haven’t, this classic trailer might help motivate them.

and what can you say, but…

Groovy.

This week in our Topics in Biotechnology course, we discussed a paper from the laboratory of Karen Vousden of the Francis Crick Institute, TIGAR, a p53-inducible regulator of glycolysis and apoptosis. I wanted to take some time to summarize the conclusions of this paper here in order to prevent any misunderstanding that might have arisen following our in-class discussion of the place of TIGAR with respect to glucose metabolism.

The body is extraordinary in its ability to maintain itself. It keeps a temperature of 37 degrees. It keeps bad stuff out and good stuff in. It has an immune system that attacks and destroys invasive viruses, bacteria, fungi, and other parasites. Cells maintain a constant pH and balance the concentrations of salts, proteins, and nutrients. All these regulatory mechanisms work together to keep us healthy and functioning properly.

One of the ways our body does this is by having our cells monitor their own health and make difficult choices when they are unhealthy. When our cells suffer damage, they work hard to repair the damage, but they also balance this against the greater good of the body as a whole. If they can repair damage, they do so, but when they can’t, cells eliminate themselves by a process called apoptosis. A central protein that controls many of these processes is the ‘guardian of the genome,’ p53.

This paper explores the role of TIGAR (Tp53-induced glycolysis and apoptosis regulator) in modulating the pro-apoptotic effects of p53 and in reducing free radicals. Specifically, TIGAR exerts its effects by rebalancing the normal metabolism of glucose during glycolysis. This prevents further damage and also allows time for repair to occur before making a decision to terminate the cell if repair is unsuccessful.

Under normal conditions, cells take up and process glucose as a fuel for making ATP, which is used directly to power enzymes and carry out all the processes that keep us alive. As part of this process, glucose is broken down stepwise during glycolysis. Some fraction of the products of this reaction gets diverted by an enzyme called PFK-2, which makes Fructose-2,6,-bisphosphate. This sugar goes on to bind to, and activate PFK-1, which keeps the pathway flowing.

Under conditions following DNA damage, p53 will become activated and the cell will arrest glycolysis as well as any cell division while it initiates DNA repair mechanisms and acts to remediate the radical oxygen species (ROSs) that are often associated with this sort of damage.

Among the many genes that are turned on to carry out these operations is TIGAR. Bensaad et al. show that the gene for TIGAR is transcribed and translated into protein, and that protein goes on to act as an enzyme to regulate metabolism.

Specifically, TIGAR functions as an enzyme with a high degree of homology to FBPase-2, which converts Fructose-2,6,-bisphosphate to Fructose-6-phosphate. This has an important regulatory function because, as stated above, Fructose-2,6,-bisphosphate is required to activate the enzyme PFK-1, which is required for glycolysis. In the absence of Fructose-2,6,-bisphosphate, PFK-1 shuts down and the products of glycolysis start backing up.

At first, this results in a backlog of Fructose-6-phosphate. As this accumulates, it will result in the accumulation of Glucose-6-phophate. With nowhere also to go, this will be processed to 6-phosphogluconolactone, making NADPH. NADPH will then oxidize glutathione, which will break down the ROS, H₂O₂ to water.

With the reduction in the number of ROSs, DNA damage will cease and repair can take place, thus diverting the cell away from a pathway leading to apoptosis.

This paper represents an amazing amount of work and is nearly bulletproof in its findings. I highly recommend it to anyone interested in how DNA damage and p53 interact with metabolic pathways and how this interaction directly leads to a more complete understanding of how p53 does its job.

Five stars.

Here is a link to an Icelandic site that is live-streaming an ongoing volcanic eruption. I just thought it was beautiful to watch geology happening in real-time and wanted to share:

https://www.ruv.is/frett/2021/03/20/beint-vefstreymi-fra-eldstodvunum

This was published on my other site, 100 films in 100 days.

Neutrophils are the first responding immune cells following exposure to pathogens.

This is due to several characteristics of these cells. Primarily, these cells are very populous in the blood. In fact, they are the most common type of circulating leukocyte, so no matter where in the body there is an insult/injury, there will be neutrophils close by to respond. Secondly, these cells are primed to respond to chemoattractants elicited by complement and from pathogens themselves.

One immediate response that will occur following an injury is the cleavage of complement molecules as they react with invading pathogens. Complement may be activated in three ways, spontaneously (i.e., the alternative pathway), by pathogen-associated carbohydrates (i.e., the lectin pathway), and by antibody (i.e., the classical pathway). In all three pathways, C3 convertase is a common element where the complement protein, C3, is cleaved into C3b, which precipitates onto the membrane of the invading cell, and C3a, which diffuses away and acts as a powerful anaphylotoxin.

C3a has a number of effects, notably activating vasculature to dilate (thus slowing blood flow in the area), upregulating adhesion molecules for cells to attach to, and promoting permeability, making it easy to Neutrophils to extravasate (leave circulation and enter the tissue). Neutrophils are quick to respond and to all of these cues and will further be attracted to the C3a gradients themselves.

Once in the vicinity of an infection, neutrophils will become more attracted by more specific attractants, such as those secreted by the pathogens themselves. One beautiful example of this kind of chemoattraction can be seen in the video of a neutrophil pursuing a bacterium in vitro:

Consider the following:

If you were given any (antibody or other) reagents you wished, how might you determine whether the neutrophil pursuing the bacterium was following a trail of fMLF? (what is fMLF? Why might this particular chemoattractant be relevant to this example? How do you determine specificity?)

This was a little hard to bear, but it’s not wrong…

a better, more complete video is this one…

A final video is…

Lastly, some important points are:

-The names and functions of the three pathways

-The initiating molecules for each pathway

-The alternative pathway is often considered a spontaneous pathway, although there are some specific activators

-IgM activates complement better than other classes of antibody

-All three pathways converge at the formation of the C3 convertase

-Complement always results in the assembly of a Membrane Attack Complex (MAC) -lyses bacteria, enveloped viruses, and nucleated cells

-C3a, 35a, and to a lesser extent C4a are anaphylotoxins

I’ve written one or two other posts about Complement including this and this.

Why does it make sense that the antibody most capable of activating complement is IgM?