A Mouse Model to Understand Lupus

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00:00:13.01 Think about the last time you had a cold.
00:00:15.08 How exactly did you know
00:00:17.10 that you were sick?
00:00:18.29 You had signs and symptoms like congestion,
00:00:21.04 a sore throat,
00:00:22.24 maybe a cough or a fever.
00:00:24.17 And what this told you was that you needed to rest,
00:00:27.00 drink a lot of fluids,
00:00:28.11 and maybe reach for that bottle of cough syrup.
00:00:30.23 But what these symptoms also tell you
00:00:33.09 is that your immune system is working really, really hard
00:00:36.14 to try to fight off that infection.
00:00:38.15 It's trying to get you healthy again.
00:00:40.16 But have you ever stopped to think about
00:00:42.21 how these cells that make up your immune system
00:00:45.03 actually know that you're sick in the first place?
00:00:48.19 I like to think of the immune system
00:00:50.16 like a security guard or a police officer
00:00:52.27 who is patrolling a building.
00:00:54.11 She's walking through the halls
00:00:55.29 and looking into various rooms,
00:00:57.13 trying to assess whether or not the building
00:00:59.26 is secure or if there's some sort of threat.
00:01:02.13 Our immune cells do the same type of thing.
00:01:05.00 They patrol our bodies,
00:01:07.20 moving through our bloodstream and into our lymph nodes
00:01:09.24 and other tissues.
00:01:11.15 And while they're at these sites,
00:01:13.12 they're looking around to see
00:01:15.17 whether or not there's some sort of infection they need to fight off.
00:01:18.21 Today, I'm gonna be talking to you
00:01:20.22 about a special type of immune cell
00:01:22.14 called a T cell.
00:01:24.03 And I think T cells are really interesting,
00:01:25.18 because the way that they search for infections
00:01:28.06 is in a very specific manner.
00:01:31.14 What I mean by this is that
00:01:33.07 we have lots of T cells in our body,
00:01:34.20 and some of these T cells
00:01:36.26 are particularly looking for certain types of infections,
00:01:39.04 like this cell here may be looking
00:01:41.18 for the virus that causes the flu,
00:01:44.04 these cells here may be looking for the bacteria
00:01:46.09 that cause strep throat, and so on.
00:01:49.07 So, let's look a little bit more in depth
00:01:51.05 at this T cell that's looking for the flu.
00:01:53.01 How exactly does it know if the flu is there?
00:01:55.27 Well, the cell has this receptor at the surface
00:01:58.13 that's shown in red.
00:01:59.28 And what it does with this receptor
00:02:02.02 is it sort of uses it as its eyes and ears.
00:02:05.02 The T cell moves around in the lymph nodes and the tissues,
00:02:08.05 and it kind of grasps around with the receptor
00:02:10.08 and samples its environment to see what's there.
00:02:13.04 And if it comes into contact
00:02:15.04 with a piece of the virus that causes the flu,
00:02:17.11 you get this sort of nice puzzle piece fit,
00:02:20.16 a perfect shape match between the bit of flu virus
00:02:23.17 and the receptor.
00:02:25.05 And this tells that T cell, there's a threat.
00:02:28.17 I need to become activated,
00:02:30.23 and I need to recruit other immune cells to this site
00:02:32.23 to get rid of this flu infection.
00:02:35.02 But if you think about it, most of the time,
00:02:37.02 we don't have the flu; we're perfectly healthy.
00:02:39.05 But our immune cells are still in this
00:02:41.26 sort of patrolling function.
00:02:43.14 They're still sampling their environment,
00:02:45.00 but what they're coming into contact with
00:02:48.01 are just totally innocuous pieces of our own cells,
00:02:50.10 our own tissues.
00:02:52.13 And so, you don't get that nice puzzle piece fit,
00:02:54.10 that sort of perfect shape match,
00:02:55.29 but you do get a little bit of interaction
00:02:58.00 of this receptor with its environment.
00:03:00.16 And so, in my research,
00:03:01.26 I'm really interested in understanding
00:03:04.12 what kind of information the cells get
00:03:07.09 from these contacts with normal, harmless things
00:03:09.18 in the environment,
00:03:11.03 and how this can go wrong
00:03:12.24 in the context of disease.
00:03:14.19 So, you could imagine that if this T cell
00:03:17.18 sees this harmless little green diamond here
00:03:19.29 and thinks that it's a threat,
00:03:21.29 even though it's not,
00:03:23.20 that cell might become activated
00:03:25.29 and start to mount an immune response.
00:03:28.09 But this would be really bad.
00:03:30.00 It would be as if this T cell is kind of sounding a false alarm
00:03:32.28 and calling in all the troops,
00:03:34.18 all the other immune cells,
00:03:36.17 to fight off an infection that isn't actually there.
00:03:39.13 And so, this is what happens
00:03:41.16 in a class of diseases called autoimmune disease,
00:03:44.16 where the T cells and other cells of our immune system
00:03:48.20 mediate attack on our bodies
00:03:50.14 even though there's no harmful or infectious agent present.
00:03:53.18 And so, there are many different types of autoimmune diseases
00:03:55.22 that can affect humans,
00:03:57.15 some of which are shown here.
00:03:59.03 And it's estimated that between
00:04:01.15 15 and 24 million people in the US alone
00:04:04.05 suffer from these and other diseases.
00:04:06.22 So, it's really important that we try to understand
00:04:08.29 what kinds of things can go wrong
00:04:11.21 with T cells and other immune cells
00:04:13.12 that can lead to this kind of disease.
00:04:16.27 And so, I've been particularly interested in lupus.
00:04:20.08 Now, lupus is a really complicated disease.
00:04:24.03 Some patients present with this sort of classic
00:04:26.12 malar butterfly-shaped rash on their face.
00:04:29.07 But not all -- it's only about 30 to maybe 60% of people.
00:04:32.18 Other patients have problems with their kidneys,
00:04:35.20 their brains, the levels of iron in their blood.
00:04:38.24 So, because lupus is such a complicated disease
00:04:40.29 and there are so many different manifestations of it
00:04:43.21 that can occur in patients,
00:04:45.25 this implies that there are probably
00:04:48.18 many different types of things that can go wrong.
00:04:50.20 So, it's important to study a lot of different types of models of lupus
00:04:55.05 to figure out all of the many different types of things
00:04:57.14 that can go wrong.
00:04:59.14 And so, for my research,
00:05:01.06 I was interested in using one of these models of lupus
00:05:02.29 to try to figure out
00:05:05.05 what is actually causing these T cells
00:05:07.02 to sound the false alarm.
00:05:08.21 Why are they starting to attack our own normal, healthy bodies?
00:05:13.07 So, it's very hard to do this kind of research on people.
00:05:16.20 So, instead, I turned to a mouse model.
00:05:19.15 Now, mice don't get that classic malar rash,
00:05:21.15 and even if they did it would probably
00:05:23.16 be pretty hard to see with all that fur.
00:05:25.28 But what we can do instead
00:05:28.03 is perform a simple blood test
00:05:30.03 to see if the mouse has a disease that looks like human lupus.
00:05:33.01 And so, in this mouse that I started studying,
00:05:35.13 we looked at its blood
00:05:37.22 and found that there were antibodies present
00:05:39.19 that can cause harm to our own tissues.
00:05:41.26 So, that meant that this mouse had lupus,
00:05:44.04 or a disease that looked like lupus.
00:05:46.21 And then, when I looked at the T cells in these mice,
00:05:49.21 they actually looked activated.
00:05:51.04 They looked like they were responding
00:05:52.26 to some sort of infection,
00:05:54.07 but the mouse was not sick at all.
00:05:55.23 It didn't have any virus or bacteria around.
00:05:59.11 And so, to try to figure out
00:06:01.07 what was going on in the T cells of this mouse
00:06:03.18 that was causing disease,
00:06:05.02 I looked at the mouse's DNA
00:06:06.25 and wanted to know whether there were any genetic changes,
00:06:09.09 any mutations,
00:06:11.07 that could be contributing to the disease.
00:06:13.10 And so, by sequencing the DNA,
00:06:15.05 I found that there was a single base pair change
00:06:18.02 in this gene called Rasgrp1.
00:06:20.19 And so, I wanted to start investigating
00:06:23.07 how altering this molecule, Rasgrp1,
00:06:25.13 could lead to these disease features
00:06:27.26 that we see in the mouse.
00:06:29.19 So, what does Rasgrp1 actually do?
00:06:32.20 Well, the gene encodes a protein
00:06:34.24 that's really important for turning signaling pathways
00:06:38.10 on inside of cells.
00:06:39.29 So, if we have our T cell here,
00:06:41.14 and let's say it's that flu specific T cell from before...
00:06:45.05 if this T cell sees a bit of the flu,
00:06:47.21 it knows it needs to become activated.
00:06:50.00 So, what is going on with Rasgrp1?
00:06:51.28 Well, Rasgrp1 activates a couple of
00:06:55.01 downstream signaling pathways,
00:06:56.19 one of which is called Ras,
00:06:58.08 and the other is called mTOR.
00:07:00.06 And for reasons that I'll explain to you later,
00:07:02.01 I became very interested in this mTOR pathway.
00:07:04.23 Now, mTOR is a kinase,
00:07:06.29 so it phosphorylates downstream proteins to turn on signaling.
00:07:10.16 And mTOR signaling is very important
00:07:12.26 for a number of different processes in T cells,
00:07:15.07 including their activation.
00:07:18.26 But let's take a step back.
00:07:20.09 This example here is showing you
00:07:22.05 what happens when that flu-specific T cell
00:07:24.17 sees and interacts with the flu virus
00:07:26.15 and becomes activated that way.
00:07:28.09 But most of the time, as you know, thankfully,
00:07:30.29 we don't have the flu.
00:07:32.13 We're not always sick.
00:07:33.24 And so, I was really interested in whether this signaling pathway
00:07:37.03 from Rasgrp1 to mTOR
00:07:38.20 was also present in cells
00:07:41.05 from just a normal and healthy mouse.
00:07:44.01 And so, to address this,
00:07:45.15 I took T cells out of normal and healthy mice,
00:07:47.29 and I measured the level of signaling
00:07:50.09 through Rasgrp1 to these pathways,
00:07:52.10 Ras and mTOR.
00:07:54.10 And what I found was that Rasgrp1
00:07:56.12 does send a low level of signal to mTOR
00:08:00.10 in these normal, healthy T cells,
00:08:02.07 but not to Ras.
00:08:03.19 So, it seemed like this mTOR pathway
00:08:05.12 was really important in the normal and healthy cells.
00:08:07.27 And what this also told me is that
00:08:10.13 T cells can experience a range of mTOR activity.
00:08:13.07 So, in the context of infection,
00:08:14.27 they get a really strong signal.
00:08:16.24 And in a normal and healthy T cell,
00:08:18.21 they get a lower level of signal,
00:08:21.02 but it's not zero -- there is still a little bit of signal.
00:08:23.20 And I think this is particularly important
00:08:25.28 for T cells to perform this sort of
00:08:28.18 police officer/security guard function.
00:08:30.16 It helps them patrol and stay alert
00:08:32.23 so that they're ready to respond and activate
00:08:35.20 really quickly when that infection does come.
00:08:39.08 So, once I established that Rasgrp1
00:08:41.18 seems to send this low-level signal
00:08:43.24 to the mTOR pathway in normal cells,
00:08:45.26 I wanted to know what was happening in the T cells
00:08:48.27 from that mouse that had lupus,
00:08:51.07 because I knew that those T cells
00:08:54.06 had a mutation in this gene, Rasgrp1.
00:08:57.07 And so, what I found when I measured
00:08:59.29 the level of mTOR signaling in these lupus T cells
00:09:02.25 was that it was actually a little bit increased
00:09:05.19 relative to a normal or healthy cell.
00:09:07.26 And so, this cell was sort of somewhere
00:09:09.28 in between the normal context and the infected.
00:09:12.19 And so, this made me really wonder
00:09:15.01 whether having this little bit of increase
00:09:17.02 in mTOR activity in the Lupus T cells
00:09:19.05 was what was actually causing the disease.
00:09:22.05 So, to address this question,
00:09:24.04 I took advantage of the fact
00:09:26.10 that there's a drug that can block, or inhibit,
00:09:28.06 the activity of mTOR.
00:09:30.02 And this drug is called rapamycin.
00:09:32.01 And actually, the word mTOR itself
00:09:34.17 stands for mechanistic or mammalian target of rapamycin.
00:09:38.01 And so, what I did is I took this drug
00:09:41.01 and I administered it to the mice
00:09:43.16 that had lupus (and this Rasgrp1 mutation)
00:09:45.24 as well as to normal, healthy, wild-type mice.
00:09:48.21 And then I measured
00:09:50.29 the level of the T cell activation
00:09:52.22 that I observed in these animals
00:09:54.14 as sort of a readout of whether they had disease or not.
00:09:57.12 So, if I just looked at the mutant mice,
00:09:59.08 without giving them any drug,
00:10:00.25 as I told you before,
00:10:02.09 there was this high level of T cell activation.
00:10:04.12 The mice kind of looked like they were responding to an infection,
00:10:06.27 even though one wasn't there.
00:10:08.14 But if I treated the mice with this drug
00:10:10.13 to turn down the mTOR signaling,
00:10:13.06 I saw a reduction in the amount of activated T cells in this mouse,
00:10:17.02 to about the levels of what you see
00:10:20.16 in a healthy, normal animal.
00:10:22.11 And so, this really showed that the lupus-like disease
00:10:25.20 in these Rasgrp1 mutant mice
00:10:27.16 was caused by that increase in mTOR signaling
00:10:29.13 that I observed.
00:10:31.17 Now, another feature of autoimmune disease
00:10:34.19 can be the accumulation of sort of specialized T cells.
00:10:38.03 And what do I mean by this?
00:10:39.16 If we think back to that security guard,
00:10:41.02 she's kind of generally walking around the building,
00:10:44.07 looking to see what's wrong.
00:10:45.23 But if she saw that there was a fire,
00:10:47.07 she would want to specifically bring in the fire department,
00:10:49.22 and not just the police force.
00:10:51.27 And so, you can think of these specialized T cells
00:10:54.00 as those firemen that would come in.
00:10:56.27 And so, if you look in a normal and healthy mouse,
00:10:59.00 you have a few of these specialized T cells.
00:11:01.25 But when I looked in the mouse with the Rasgrp1 mutation,
00:11:04.26 I found that there were actually
00:11:07.06 a lot of these specialized T cells,
00:11:09.01 which is depicted in this graph here.
00:11:10.18 And if I administered that drug, rapamycin,
00:11:13.00 to turn down the mTOR signaling,
00:11:15.20 I found that this population of specialized T cells decreased.
00:11:20.07 So, it showed, again, that these specialized T cells
00:11:23.00 were accumulating in a manner that was dependent
00:11:25.08 on having some mTOR signaling in the cells.
00:11:30.23 And so, now I've shown you
00:11:32.24 that there is this signaling pathway
00:11:34.18 from Rasgrp1 to mTOR
00:11:36.10 that seems to be increased
00:11:38.11 when you get this single nucleotide change in Rasgrp1.
00:11:41.10 And this leads to a lupus-like disease in the mice.
00:11:44.14 Now, I told you that mTOR signaling controls T cell activation
00:11:48.15 and that I saw that the T cells
00:11:50.15 in these Rasgrp1 mutant mice
00:11:52.07 looked like they were activated.
00:11:54.00 And I also saw an accumulation
00:11:55.23 of these specialized T cells
00:11:57.19 that may be due to increased proliferation,
00:11:59.19 which is also a process driven by mTOR.
00:12:02.20 But I wanted to know from here,
00:12:04.09 what might be going on inside the cell,
00:12:06.14 at the molecular level,
00:12:08.08 that could be causing the T cells to become activated?
00:12:12.04 So, not just the signaling,
00:12:13.24 but what was sort of connecting the signaling to the activation.
00:12:17.17 And so, from here, I started to look
00:12:19.26 at this process of translation
00:12:21.25 -- so, going from mRNA into protein --
00:12:23.29 because this is a process that's known to be
00:12:26.27 very tightly controlled and regulated
00:12:28.17 by mTOR signaling.
00:12:31.21 So, again, translation
00:12:33.09 is this process of going from mRNA into protein.
00:12:37.02 And to see whether translation
00:12:39.22 might be altered in my Rasgrp1 mutant mice,
00:12:42.06 I started by looking at one single gene, called Gata3,
00:12:46.10 as a proof of concept.
00:12:48.11 Now, Gata3 is known to be very important
00:12:50.15 for the regulation and development
00:12:52.19 of these specialized kind of firefighter T cells,
00:12:55.25 and so that's why I chose to look at it first.
00:12:58.13 So, to see if Gata3 was regulated
00:13:01.10 at the level of translation,
00:13:03.00 I looked at its levels of mRNA
00:13:05.18 in just a normal, healthy, wild type mouse
00:13:07.11 compared to the Rasgrp1 mutant.
00:13:09.03 And I didn't really see any changes.
00:13:11.09 So, it looked like they had about the same level of mRNA.
00:13:14.23 But when I measured the protein levels of Gata3,
00:13:17.14 I found that the mutant T cells
00:13:20.02 had about twice as much protein,
00:13:22.15 so it seemed like there was something happening
00:13:24.08 between this step of mRNA to protein
00:13:26.10 to cause that to accumulate,
00:13:28.02 and so I thought this might be due
00:13:30.15 to an increase in translation in the mutant T cells.
00:13:33.11 Now, this data was really exciting,
00:13:35.03 and it made me want to look at more and more genes.
00:13:37.04 But instead of just picking ones that made sense to me,
00:13:39.09 I wanted to take a more unbiased approach
00:13:41.18 and instead look at all of the genes
00:13:43.22 in the genome.
00:13:45.10 And so, to do this,
00:13:47.16 I used a really cool new technology.
00:13:49.12 The first step was to just isolate
00:13:52.01 all of the mRNA from T cells,
00:13:53.23 to get a representation of which genes
00:13:55.28 were expressed and at what levels.
00:13:57.25 And this is called RNA sequencing.
00:14:00.21 But to look specifically at the mRNAs
00:14:02.26 that are being translated,
00:14:05.00 what I did was a technique called ribosome profiling.
00:14:08.01 So, I took the ribosomes out of cells
00:14:10.14 and isolated the mRNAs
00:14:13.03 that are bound to those ribosomes
00:14:14.18 -- so, only those mRNAs that are actively
00:14:17.09 being translated in the cells --
00:14:18.24 and then I sequenced that RNA as well.
00:14:21.17 And by comparing the level of expression
00:14:23.29 in the ribosome data set to the mRNA,
00:14:27.02 I could get a sense of which genes
00:14:29.15 were being regulated at the level of translation.
00:14:31.29 And I was really excited to see
00:14:35.06 that there were around 600 genes
00:14:37.13 that seemed to have increased translation
00:14:39.13 in the mutant T cells compared
00:14:41.24 to the normal, wild type ones.
00:14:43.27 And this is a really ongoing, active area of research
00:14:46.25 in my lab now,
00:14:48.20 to try to figure out what these genes are doing in the T cells
00:14:52.12 and how they may be contributing to the disease.
00:14:54.10 And so, this is an exciting, open question
00:14:56.20 that we're hoping to answer later on.
00:15:00.21 So, by studying this Rasgrp1 mutant mouse,
00:15:03.07 I was able to learn a lot of different things
00:15:05.04 about T cell biology
00:15:07.17 and how it relates to disease.
00:15:09.08 I showed you that T cells
00:15:11.18 have this sort of range of mTOR activity,
00:15:13.12 and that normal, healthy T cells
00:15:16.01 have a little bit of mTOR signaling
00:15:18.03 that helps them perform this patrolling function --
00:15:21.27 it helps them stay alert and ready to respond.
00:15:24.15 But if you have a mutation in this gene, Rasgrp1,
00:15:28.05 and it increases the level of mTOR signaling a little bit,
00:15:32.16 this can cause that T cell to increase its mTOR activity
00:15:36.24 and then that T cell can become more activated,
00:15:39.20 can start to become more specialized,
00:15:41.24 and can... star...
00:15:44.10 can contribute to the lupus-like disease that we see in the mice.
00:15:47.04 And we think that this is due to mTOR's regulation
00:15:50.18 of translation in cells.
00:15:53.20 And so, this is what's sort of causing
00:15:57.00 the T cells in this model
00:15:59.02 to sound that false alarm.
00:16:00.19 They have this little bit of elevated mTOR activity
00:16:02.20 and they're... instead of knowing that everything is fine,
00:16:06.16 they are contributing to disease.
00:16:09.01 And so, this research, I think,
00:16:13.11 leads to a lot of really interesting questions
00:16:15.03 and future directions.
00:16:16.23 So, is this phenomenon of increasing the level of mTOR activity
00:16:19.26 in a T cell
00:16:21.23 only applicable to this model of lupus,
00:16:23.19 or is it true for other types of autoimmune diseases as well?
00:16:26.25 Is this sort of a universal,
00:16:28.20 or a unique, phenomenon?
00:16:30.02 Additionally, I looked at the mTOR pathway
00:16:33.09 in these T cells from these lupus mice
00:16:36.06 and saw differences,
00:16:37.29 but perhaps other signaling pathways are also altered
00:16:40.19 in normal T cells in the context of autoimmune disease.
00:16:44.10 And perhaps these other pathways
00:16:46.14 also sort of tune the reactivity of those cells.
00:16:49.23 And in general, I think this research
00:16:52.08 shows that performing basic research
00:16:54.17 to understand signaling pathways
00:16:57.08 and general T cell biology
00:16:58.25 is a way to get a deeper understanding
00:17:01.06 of autoimmune diseases like lupus,
00:17:03.01 and hopefully one day will contribute
00:17:05.08 to our ability to design
00:17:08.05 better, more rational therapies to help patients.

This Talk

Speaker: Darienne Myers

Audience:

Educators of H. School / Intro Undergrad
Student
Educators of Adv. Undergrad / Grad
Researcher
Educators

Recorded: May 2018

Talk Overview

Lupus is a disease in which the immune system becomes erroneously activated and attacks a patient’s own tissues. Using a mouse model of lupus called Rasgrp1, Dr. Darienne Myers investigates the molecular mechanisms of this autoimmune disease. Specifically, she asks how the immune system knows when to mount a response. She discovers increased levels of mTOR (Target of Rapamycin) signaling and protein translation in the Rasgrp1 mouse’s T cells, which contributes to the lupus-like disease displayed by the animal.

This talk is part of the Young Scientist Seminars, a video series produced that features young scientists giving talks about their research and discoveries.

Speaker Bio

Darienne Myers

Darienne Myers received her Bachelor’s Degree in Biology from Amherst College. After this, she worked as a research assistant in Dr. Fred Alt’s lab in Boston, MA, where she first became excited by Immunology research. Darienne then pursued a Ph.D. in Biomedical Sciences with a focus in Immunology at the University of California, San Francisco… Continue Reading