Tag Archives: Autoimmune disease

The proteasome branches out

The surface of a protein is not at all like a ball of yarn, even though they are both one long string. This has profound implications for the immune system. Look at any solved protein structure. The backbone bobs and weaves taking water hating (hydrophobic) amino acids into the center of the protein, and putting water loving (hydrophilic) amino acids on the surface. So even though the peptide backbone is continuous, only discontinuous patches of it are displayed on the protein surface.

Which is a big problem for the immune system which wants to recognize the surface of the protein (which is all it first gets to see with an invading bug). Now we know that foreign proteins are ingested by the cell, chopped up by the proteasome, and fragments loaded on to immune molecules (class I Major Histocompatibility Complex antigens) and displayed on the cell surface so the immune system can learn what it looks like and react to it. The peptides aren’t very long — under 11 or so amino acids, but they are continuous.

What if the really distinct part of the protein surface (e.g. the immunogen)  is made of two distinct patches from the backbone? A fascinating paper shows how the immune system might still recognize it. Chop the protein up into fragments by the proteasome, and then have the fragments from adjacent patches put back together. You know that any enzyme can be run in reverse, so if the proteasome can split peptide bonds apart it can also join them together.

This is exactly what was found in a recent paper — Science vol. 354 pp. 354 – 358 ’16. The small peptides (containing at most 11 amino acids) finding their way to the cell surface were analyzed in a technical tour de force. In aggregate they go by the fancy name of immunopeptidome. They found that the proteasome IS actually splicing peptide fragments together. This is called Proteasome Catalyzed Peptide Splicing (PCPS). The present work shows that it accounts for 1/3 of the class I immunopeptidome in terms of diversity and 1/4 in terms of abundance. One-third of self antigens are represented on the cell surface of the immune cell line they studied (GR-LCL the GR-lymphoblastoid cell line) ONLY by spliced peptides. The ordering of the spliced peptide was the same as the parent protein in only half. There was no preference for the length of the protein skipped by the splice.

The work has huge implications for immunology, not least autoimmune disease.

So today I wrote the author the following

Dr. Mishto

Terrific paper ! Do you have any evidence for the spliced peptides being spatially contiguous on the surface of the parent protein. Have you looked?

This makes a lot of sense, because the immune system should ‘want’ to recognize protein conformations as they exist in the living cell, rather than stretches of amino acid sequence in the parent protein. Also, with few exceptions the surface of a given protein in vivo is a collection of discontinuous peptide sequences of the parent protein. I’ve always wondered how the immune system did this, and perhaps your paper explains things.

Luysii

and got this back almost immediately

Dear Luysii

Interesting idea. We shall have a look for few examples where the crystallography structure or the parental protein is disclosed already.

regards

Michele

It doesn’t get any better than this. Tomorrow I will be exactly 78 years and 6 months old. It shows I can still think (on occasion).

Addendum 17 Nov ’16;  It looks as though proteins are fed into the central cavity of the proteasome as a completely denatured single strand.  See figure 5 of PNAS 113 pp 12991 -m12996 ’16.  The channel to get in appears quite narrow.

Is a rational treatment for Multiple Sclerosis in our future?

Two very recent papers taken together point the way to a rational treatment of multiple sclerosis (and probably all autoimmune disease). The short story:
Paper #1 found a way to find the antigen or antigens patients with MS are reacting to
Paper #2 found a way to selectively impair the response to an inciting antigen without clobbering the whole immune system

Some history: Some evening in 1966 or 1967 a fellow neurology resident and I were sitting on the ward having dealt with the complications of high doses corticosteroids for a case of optic neuritis (often the first sign of MS). I said, some day they’ll look at what we’re doing the way we look at docs of 200 years ago using leeches (and bloodletting). As a kid, I remember my parents driving into Philly. Shortly after getting over the Ben Franklin bridge we’d pass a pharmacy offering leeches on its sign.

It was obvious even back then that MS in some way was an attack by the immune system on the brain. Finding the particular antigen the system was reacting to would lead us to the cause and hopefully less simplistic treatment than clobbering the immune system. We didn’t know all the proteins we had or even how many, so people would look for antibodies to a variety causes (which they’d arrived at by reasoning, not data). Increased antibody titers to a variety of viruses were found, but that led nowhere. No one ever isolated a virus from MS brain, although sightings on electron microscopy were eagerly reported. Eventually it became obvious that the immune system was on high alert with increased antibodies to lots of things.

This leads to paper #1 [ Proc. Natl. Acad. Sci. vol. 113 pp. 2188 – 2193 ’16 ] To make a long story short they used something called the Human Protein Atlas Program to find what proteins the antibodies in MS patients were reacting to. So rather than having a theory about what MS patients might be reacting to and testing it, they looked at all proteins and watched. It’s the difference between being a Greek philosopher reasoning things out from first principles and collecting data. Only when the technology is available can you stop a priori theorizing and just look. Don’t be too hard on the earlier researchers, they didn’t have the tools.

The found that MS patients were reacting to a protein called anoctamin2, which actually showed increased expression near and inside the demyelinating plaques of MS.

For the gory details keep reading, otherwise skip to **** where I’ll discuss paper #2

Gory details — The Human Protein Atlas produces human protein fragments, selected on the basis of their low similarity to other proteins in the proteome. [ Science vol. 347 1260419 (23 Jan) ’15 ] The atlas hopes to find out where and how much of each our proteins is at the tissue and cellular level. It is based on antibody based profiling on tissue microarrays (of proteins?). This based on transcript expression (RNA-Seq), and immunohistochemistry (24,028 antibodies coresponding to 16,975 protein coding genes). 44 tissues were studied. The antibodies produced more than 13 million tissue based mmunohistochemistry images. They also report subproteomes (secreted proteins n = 3,171, and membrane bound proteins n = 5,570). Interstingly there was an overall concurrence between mRNA and protein levels for a given gene product across various tissues.

The PNAS paper profiled 2,169 plasma samples from MS cases and population based controls (with neurologic disease) using bead arrays built with 384 human protein fragments seleted from an initial screening with 11,520 antigens. There was increased reactivity to anoctamin2 (aka TMEM16B) in MS vs. controls (by how much?). This was corroborated in independent assay with alternative protein constructs and by epitope mapping with peptides covering the identified region of anoctamin 2.

ImmunoFLuorescence in human MS brain tissue showed increased anoctamin2 expression as small cellular aggregates near and inside MS lesions. The controls had other neurologic disease. There was a 5.3 fold change in fluorescence intensity in the MS group. The antibodies are directed against the amino terminal region.

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Paper #2 — [ Nature vol. 530 pp. 422 – 423, 434 – 440 ’16 ] basically found a way to knock out the immune system’s response to a single antigen — not all of them. The point is that just an antigen by itself isn’t enough to turn the immune system on. A costimulatory molecule must also be present on the antigen presenting cell. If it isn’t there the immune system is actually turned off by forming regulatory T cells (which even though they are part of the immune system they actually turn it off).

One can form models of human autoimmune disease in mice. Two such are EAE (Experimental Allergic Encephalomyelitis) formed by giving the animal myelin basic protein (a constituent of myelin which is attacked in MS), and rheumatoid arthritis (formed by giving collagen to the animals). What is so great about this paper is that MHC II carrying peptides from collagen suppress disease in a mouse model of rheumatoid arthritis, but NOT in mice with EAE. MHC-II carrying CNS antigen peptides control EAE but not collagen induced arthritis.. In addition neither treatment impaired the immune response to infection — something that almost always happens when you clobber the immune system.

Well it’s a long way from the lab to the bedside, but imagine finding what the immune system is reacting to and stopping it (without stopping the immune system). That’s what these two papers portend. Exciting times.