A Guide to Chapter Five of "Chronic Fatigue Syndrome A Biological Approach' (Edited by Patrick Englebienne Ph.D., Kenny DeMeirleir M.D, Ph.D., CRC Press. Washington D.C. 2002) by Cort Johnson

Chapter Five: The 2-5A Pathway and Signal Transduction: A Possible Link to Immune Disregulation and Fatigue By Patrick Englebienne, C. Vincent Herst, Marc Fremont, Thierry Verbinnen, Michel Verhas and Kenny De MeirLeir  

Chapter five is EXTREMELY difficult.  A good portion of it is spent simply explaining how the signal transduction pathways work. The authors suggest that many of the symptoms found in CFS could be explained by improper signal processing.  First the types of receptors are discussed, then the signaling pathways that are activated by the receptors, and finally the object of interest in CFS – the interferon signaling pathway and its connection with RNase L. 

INTRODUCTION - Signal transduction (the processing of an external or internal signal via a receptor found on a cellular membrane) involves a complex chain of biochemical interactions (a cascade) which ultimately results in gene activation in the nucleus. 

The activation or inactivation of every step in the signal transduction chain is accomplished through phosphorylation and dephosphorylation (the addition or removal of a phosphate group from a protein). As such the phosphorylating and dephosphorylating enzymes (kinases, phosphatases) are of paramount importance. \

Phosphorylation involves replacing a hydroxyl group (OH) with a phosphate group in certain proteins. The strong negative charge that phosphate groups carry allows them to change the shape of a protein by attracting positively charged and repelling negatively charged amino acid groups.  Changes in a protein shape uncovers previously hidden catalytic sites and allows the protein to become activated, or bind with another protein or ion.

Although these enzymes function in a wide variety of signaling pathways they have broad specificities; that is they interact with a broad variety of substrates. In order to achieve the specificity needed to run a complex system several methods are used; enzymes are combined together to create unique signals, and special signaling molecules or dormant enzymes are embedded deep within the signaling network. 

The ‘cross-talking’ that takes place between some signaling pathways increases the systems complexity.  Some pathways interact synergistically to activate gene transcription and others produce signaling molecules which interact with other cells.  To make matters worse, the same ligand (receptor binder) can produce diametrically opposite effects when it binds to different receptors. 

The complexity of the cellular signaling pathway system underscores how regulatory failures in one part of the system can potentially have negative consequences in another.

THE SIGNAL TRANSDUCTION SYSTEM

Signal transduction begins when a ligand (a ‘key’ that fits into the receptors ‘lock’) binds with a receptor and initiates a series of events that can range from the cell shutting down its ion channels, to rearranging its internal structure, to creating a new protein to do a task.  The receptor can occur on the external cell membrane, on the cellular membranes surrounding the organelles in the cell, in the cytosol or in the nucleus.    The receptors are the sensory system of the cell; they provide cells with signals that tell them how to behave appropriately in their environment.  Disruption of the signaling pathway means a dysfunctional cell, and ultimately, a dysfunctional body).   

Receptors inside the cell belong to the steroid and thyroid hormone receptors superfamily. Upon activation they bind with a ‘responsive’ DNA element (a hormone responsive element or HRE) and activate the transcription of a gene. Transcription occurs when messenger RNA (mRNA) is produced. Messenger RNA contains the directions needed to make proteins; upon leaving the nucleus it goes to the ribosomes where the proteins it codes for are made. 

Three types of receptors are found on the cells external membrane; intrinsic activity receptors, g-protein coupled receptors, and those associated with channels.  Receptors associated with channels are beyond the scope of this chapter.  Intrinsic activity receptors can, upon activation, phosphorylate themselves using their own kinase enzymes. Most `receptors are activated by a phosphate unit, they then activate a separate kinase which advances the signaling cascade.  Insulin and growth factor receptors are intrinsic activity receptors, as are the receptors for lymphocyte activation.

G-protein receptors make up a superfamily of receptors that play important roles in pathways involving neurotransmitters, immune recognition, hormones and the sensory system.  As noted above, these proteins bind to the guanine triphosphate (i.e. GTP binding or G-proteins) sections of amino acids.  When they bind to guanine nucleotides, they activate GDP by converting it to GTP which is then able to bind to an enzyme or ion channel and activate it.   This process is mediated by adenylate cyclase. Adenylate cyclase converts ATP to ADP and generates cAMP (cyclic adenosine monophosphate). Cyclic AMP activates protein kinase A, which then phosphorylates other members of the cascade). 

THE SIGNAL TRANSDUCTION CASCADES

The five different signaling cascades described in this section are all induced by interferons. They give an indication of the far reaching effects that the IFN dysregulation found in CFS may have.  This section, too, is quite technical.

There are several pathways incorporated in the signal \ transduction cascades.  The Ras/Rho and PI-3K are probably the best known.  Ras and PI-3K proteins play a major role in the signal transduction of growth factor receptors.  Rho proteins, which are homologous to Ras proteins, are G-proteins which regulate the actin cytoskeleton.

These activities may seem ‘obscure’ but they are both quite fundamental and are of interest in CFS.  Growth factors stimulate the growth of other cells. Many hormones, including androgens and estrogens (sex hormones), growth hormone, prolactin (stimulates lactation), and thyroid hormone, are growth factors as are some vitamins, interleukins, etc. Actin is a protein found especially in the microfilaments that make up the cellular skeleton.  It is involved in, among other things, antigen presentation and apoptosis (cell suicide). Without proper signal processing cells would be impervious to hormones, vitamins, or be able to respond to invaders or engage in apoptosis (i.e. defend themselves from pathogens) properly. 

The Ras pathway occurs parallel to and in conjunction with a pathway (Rac) that is stimulated by cellular stress. We shall see that these researchers believe cellular stress is an important ‘predisposing factor’ in CFS.  These inter-related signaling pathways activate a variety of kinases (MAPKS, ERKS, MKK, JNKS, etc) whose phosphorylating activities ultimately cause nuclear transcription (and the production of mRNA) to take place.

 These signaling cascades are called the mitogen activated protein kinase cascade.  Mitogens are substances, which stimulate cell mitosis or cell division.  Both growth factors and mitogens stimulate the Ras/PI-3K signaling cascades. They are described in greater detail in this chapter than will be given here. A few general comments will have to suffice.

The different sections of this mitogen activated protein kinase (MAPK) cascade activate transcription factors such as NF-kB that induce the transcription of genes that produce such important immune factors as interferon B, cox-2, and nitric oxide synthetase (iNOS) and heat shock proteins. The activities these substances are engaged in are examined with some detail.

Besides stimulating the production of RNase L, interferon B or IFN-b stimulates the expression of HLA class I and II genes, activates cytotoxic T-cells and (to a lesser extent) macrophages and NK cells, and inhibits cell growth.  HLA molecules are what antigens presenting cells use to present viral peptides to T and B cells for examination.  Essentially IFN-b stimulates a wide variety of cells to turn on their ‘radar’ and be on the alert for foreign invaders.  Inhibition of IFN-b would allow invaders to slip past the immune systems defenses.

Nitric oxide synthetase (iNOS) catalyses the production of nitric oxide.  Nitric oxide (NO) is a highly reactive molecule involved in a host of functions, including smooth muscle relaxation and vasodilation, neuro-synaptic regulation, and in the immune system, macrophage activation and apoptosis.  Macrophages use NO to kill intracellular parasites.  Natural killer cell activity may be disrupted in CFS via a nitric oxide mediated pathway.  Nitric oxide reacts with superoxide to form an even more reactive molecule, peroxynitrate.  Nitric oxide dysregulation is believed by Martin Pall to play a major role in the pathogenesis of CFS, FMS, and MCS.  It has been implicated in a host of diseases.

Cox II is an enzyme that makes prostaglandins and is involved in the inflammatory response.  Prostaglandins dilate blood vessels and increase vascular permeability, pain sensitivity and cause fever.)    

The PI-3K (phosphatidylinositol 3’-kinase pathway) activates (phosphorylates) protein kinase B (PKB) which in turn phosphorylates a wide variety of substances that participate in a wide variety of metabolic activities essential to cellular energy supply and protein synthesis.  These include glucose, glycolysis and glycogen uptake.  (Breaking down glucose through glycolysis results in the production of ATP.  Glucose is converted into glycogen for storage in the tissues.)  

The PI-3K pathway is also fundamentally important in regulating cell apoptosis. (Cell apoptosis or programmed cell death will be looked at closely in this text.   A suicide program invoked in damaged or virally infected cells results in the cell degrading its proteins, chopping up its nucleus and then fragmenting itself into membrane bound pieces.) The PI-3K pathway inhibits the production of caspases (proteolytic enzymes) and proteins that both prevent and enhance apoptosis.

INTERFERON RECEPTORS AND SIGNAL PROCESSING

This very difficult, and to me, sometimes almost incomprehensible section describes how the interferon signaling pathway works, and the role that IFN dysregulation might have on PKR (protein kinase R) production, viral protection, and cell apoptosis. STAT I, an important component of IFN induced gene transcription, appears to be of special consequence in CFS.  There is a summary at the end!

Interferons modulate the immune response and induce a antiviral state upon activation. IFN’s a,b (type I IFN’s) are secreted when a cell is infected with a virus.  So far 14 different IFN-a subtypes that elicit different responses have been identified.  IFN-y is secreted by T-cells upon activation by natural killer (NK) cells.

The signals that both type I and II IFN’s produce both go through the STAT proteins. The STATS are the last processing agents the signal passes through prior to its entering the nucleus. STAT I, in particular, plays a critical role in mediating Type I and II IFN activity.  STAT I expression was examined in the PBMC’s of CFS patients to determine if the dysregulation found in the IFN pathways in CFS extended that far. These researchers found that as the levels of the 37k-Da RNase L fragment rose, STAT I protein levels diminished until, at the higher levels of 37-kDa, the STAT I proteins were completely degraded.  This suggests that whatever degrades RNase L not only degrades RLI (RNase L inhibitor) but also degrades STAT - the protein responsible for switching on the interferon producing genes in the nucleus.  STAT I degradation could, therefore, explain the lack of responsiveness to IFN’s found in CFS.

This indicates that the IFN signal produced by virally infected or otherwise compromised cells may simply not getting through to the nucleus.  Thus the inhibition of the Th1 side of the immune system may arise simply because the T-cells are not getting the message. What starts as a disruption in the viral defense system becomes a far larger issue when some of the key messengers of the immune system, the interferons, are effected.  Remember that over 100 interferon stimulated genes has thus far bee identified. Contrary to what is stated above, however, STATS are not the only signaling proteins that the interferon signal is processed through.

The authors have shown us how the signal for interferon production gets to the nucleus. Now they look at what happens to it in the nucleus. The mechanisms used in the nucleus of the cell to generate IFN expression are less clear. We know that activation of the interferon regulating factors 3, 7 (IRF’s 3,7) is required but how these IRF's are activated is unclear.  PKR induction by dsRNA was believed, at one point, to lead to IRF phosphorylation but recent evidence indicates that a new cellular kinase may be responsible. 

We do know that cellular stress can induce the MAPK signaling pathway to activate IRF-3. We will see that cellular stress or viral activity appears to jumpstart the process leading to CFS.  Interestingly, the phosphorylating sequences in the IRF’s are different depending if they activated by cellular stress or viral attack. IRF activity following viral activation is far more extensive and includes nuclear translocation (transfer of a section of one chromosome to a non-homologous (i.e. different) chromosome) and transactivation (?). IRF activation via psychological stress does not. This suggests that, contrary to widely held beliefs that all stressor are equal, that they are not; viral stressors prompt far more IFN activity in a cell that do psychological stressors.

Type II interferons (IFN-y) are produced during T-cell activation and by NK cells. IFN-y is one of the ‘genes’ regulated by Type I IFN’s.  This means that the gene disruption in the STAT-1 portion of the IFN I signaling pathway could have profound effects on IFN-y activity and therefore the production of Th1 type (pro-inflammatory) cytokines. STAT4 is part of the signaling pathway leading to IL-12 production; since IL-12 signals for IFN-y production inhibited STAT 4 activity could result in reduced IFN-y production. The problems this could cause are seen below.

A Guide to Type II Interferons - Upon contact with an infected or damaged cell macrophages secrete IL-1 and 12. When the precursors of T cells come across macrophages or other APC’s, they search its surface to see if it displays a harmful antigen.  If it does, and the T-cell has already been  primed by the presence of IL-12, then the T-cell becomes activated, grows, and then splits apart into several T-cells and begins secreting IFN-y and TNF-a.  These cytokines prompt the macrophages to seek out and destroy the invaders. The IFN-y produced acts to, among other things, suppress the development of Th2 cells. The antagonistic nature of the THI/Th2 interaction means that stimulation of one side results in inhibition of the other. 

IFN-y figures in a very wide range of activities. It is of particular interest in CFS because it is central to activation of that arm of the immune system (Th1) that appears to be inhibited in CFS. IFN-y ramps up the alert level of antigen presenting cells by enhancing the expression of the (MHC) molecules they use to display foreign antigens on.  Antigens are anything - viruses, bacteria, and chemicals - that provoke an immune response.  IFN-y also activates cytotoxic T-cells (Tc) (which kill virally infected cells), increases natural killer (NK) cell activity, and activates monocytes/macrophages (the sentries of immune system). IFN-y differentiation of dendritic cells - the major antigen presenting cells - triggers the adaptive immune response (ThI) response which may be deficient in CFS.

IFN-y, among others, is the one of the immune system mediators NK cells produce. The poor NK cell function found in CFS could, therefore, result in low ThI differentiation. 

So how to explain high IFN-y levels but normal IL-12 levels in CFS patients?  It may be that the STAT 4 signal transducers are working properly (STAT 4 activates both IL-12/IFN y) but that STAT 1 (which regulates IFN-y) is dysfunctional.  (STAT 1 induces NF-kB which inhibits inflammation.  High IFN-y production would seem, however, to be paradoxical in CFS since it is the Th1 not the Th2 system in the immune system that appears to be depressed.  This conundrum will be explained in Chapter eight).  The authors suggest that dysregulated IFN-y production by type I IFN’s in monocytes might lead to high IFN-y levels. On the other hand reduced IL-12 production may result from increased sensitivity to glucocorticoids.  Combined, this all results in poor NK activity.

 The pituitary and hypothalamus respond to stimulation by IL-1, IL-6 and TNF-a (all from macrophages) by secreting adrenocorticotropin hormone (ACTH).  ACTH stimulates adrenal glands to secrete corticosteroids (glucocorticoids).  A rapid rise in corticosteroids is often observed during the early phases of infection.  Interestingly enough, corticosteroids down regulate IL-1 synthesis, and are part of a negative feedback loop between the nervous and immune systems.  High sensitivity to glucocorticoids might result in increased down regulation of IL-1 synthesis and this could, by inhibiting macrophage functioning, conceivably result in reduced IL-12 production??? Because NK cells are activated by IL-12, reduced 1L-12 production would then result in poor NK cell functioning.

The authors suggest that STAT 4 signaling pathway - which plays a major role Th1 differentiation – is operating normally.  STAT I, however, which regulates both the IFN I (virus activated) and II (T-cell, NK cell activated) systems, appears to be dysfunctional in CFS.  This may have consequences beyond those concerning immune defense. The genes regulated by type I and II IFN’s also play a major role in cell suicide - the process by which infected or damaged cells remove themselves before they cause too much trouble.

Type I IFN’s enhance the signaling cascade by increasing the expression of an interferon regulatory factor (IRF) and of several antiviral genes coding for a variety of effector proteins. (Effector proteins control protein synthesis at the genetic level.)   When PKR is inactive it binds with STAT I and inhibits type I IFN activity.  The authors suggest that both the upregulation in PKR and the improper induction of 2-5OAS found in CFS could be caused by the STAT I degradation observed in CFS. (This is a new twist.  If STAT I is degraded does PKR have nothing to bind to and so become upregulated?  Are the authors suggesting that upregulated PKR results in upregulated apoptosis, and this increased apoptosis produces the small RNA fragments that improperly activate 2-5OAS?).

Several other antiviral proteins are activated by Type I IFN’s. (This section takes a look at the problems that proteins effected by a dysfunctional IFN I signaling pathway could cause in CFS). The MxA protein inhibits RNA virus multiplication.  Over expression of MxA induces apoptosis. (This could occur if an upregulated PKR enzyme induced an over expression of Type I IFN’S (?))  MxA operates differently in the cytoplasm and in the nucleus.  In the cytoplasm it stops viral proteins from entering the nucleus (where they would insert themselves into the DNA and induce the cell produce substances resulting in viral replication).  In the nucleus MxA stops the virus from engaging polymerase activity.  (The virus is stopped from building blocks of viral DNA.  Down regulation of this protein could obviously result in increased viral activity in CFS.  Upregulation would result in increased apoptosis. Which is happening in CFS? I have no idea?).  

Ubiquitin, a protein that participates in the genes coding for cell apoptosis (by targeting the appropriate proteins for destruction), and is involved in inducing NK cell proliferation and activity, is induced by Type I IFN’s. (Right away we have a possible reason for low NK cell activity and inhibited apoptosis.  Low IFN signal transmission could result in low NK cell activity and inhibited apoptosis.)

P53 represses the gene coding for p202, a protein which negatively regulates apoptosis induced by p53.  Degradation of p53 has been observed in CFS and might be responsible for the upregulation of p202 and a subsequent apoptotic inhibition.

Type I IFN’s (IFN a’s) also enhance the Fas ‘death receptors’ on PBMC’s and T-cells.  Upregulated IFN a’s could result in increased apoptosis in monocytes/macrophages.

NF-kB is a nuclear transcription factor that plays an integral role in mediating the responses to the genes stimulated by IFN’s.  NF-kB is essential for nitric oxide synthetase production by macrophages.  (Macrophages kill intracellular parasites using nitric oxide and other substances).   NF-kB is also involved in apoptosis via its transcription of Bcl-2.  Lastly PKR is involved in NF-kB activation through its phosphorylation of the I-kB inhibitor and may also figure in a separate NF-kB signaling pathway that was recently discovered. (As mentioned earlier PKR upregulation is often found in CFS.  We will learn in the next chapter that NF-kB inhibitor is also fragmented in CFS.  An upregulated PKR system could apparently result in increased iNOS production, increased prostaglandins and cox levels and reduced apoptosis).

Apoptotic signals induced by the Type I IFN pathway are enhanced by several members (retinoic acid, tamoxifen) of the thyroid and steroid receptor superfamily.    The effects of these proapoptotic signals are mediated by several genes associated with ‘retinoic acid IFN-induced mortality (GRIM).  One product of one of these genes (GRIM 12) is an enzyme (thioredoxin reductase) which reduces p53, thus enabling its interactions with DNA. Another protein (GRIM 19) activates caspase 9.  The inactivation of p53 and caspase 9 inactivation observed in PBMC’s of CFS patients, might result in disregulation of the apoptotic balance of type I IFN’s. 

A Summary: What do we know after this most difficult of sections? (That we are confused, certainly.) We know that STAT I, a very important transcription factor that regulates gene expression of the type I and II  IFN’s, is degraded and may block IFN induced activity. IFN-y production itself  does not appear to be inhibited but the authors believe that cells in CFS patients are not responding to it because its signal is blocked by STAT I degradation. Because IFN-y is intimately involved in the adaptive Th1 response, this could inhibit the bodies ability to respond to intracellular invaders.

The increased sensitivity to glucocorticoids found in CFS down regulates the production of a major cytokine, IL-1, that is important for macrophage functioning.  Since macrophages activate NK cells, this could lead to reduced NK cell activity.  

Because both IFN types pay a role in regulating apoptosis, the problems spread to the cell suicide program - an essential component of the immune response (see next chapter).  Not only that but the authors suggest that STAT I disruption could be responsible for the PKR upregulation and improper 2-50AS activity found in CFS!  RNase L disruption could also originate in STAT 1 degradation.  RNase L and PKR are both components of the IFN pathway that leads through the STAT proteins. If I'm reading this right then the degradation of the STAT I protein could be central to the problems in CFS.

TYPE I INTERFERON-STIMULATED GENES AND THE THYROID RECEPTOR

The 2-5 synthetase enzymes are activated by single stranded and double stranded RNA’s. The 2-5A synthetase family includes a variety of enzymes of different weights (44/46, 69/71, p100). The enzymes catalytic abilities - their ability to synthesize ATP into 2-5A fragments - is highly dependent on their weight. The p100 isoforms produce more 2-5A dimers - which do not activate RNase L - and the p69 isoforms produce more 2-5A trimers - which co-activate RNase L. 

Type I IFN’s also induce three proteins called 2-5 oligoadenylate synthetase-like protein (2-5OASL) that are, not surprisingly given their name, closely related to 2-5OAS. An analysis of the amino acid sequence of all six of these 6 proteins indicated that they are least similar in the N-terminal region where the catalytic activity of 2-5 OAS+ is located. (It appears that the 2-5OASL’s may be able to bind to ATP but cannot cleave it- they cannot produce 2-5A).

A ‘blast search’ of p59SOASL revealed that it was a thyroid receptor (TR) interacting protein (a TRIP 14).  TRIPS interact with thyroid and retinoid receptors. Because OASL proteins are found in the nucleus (as well as the cytoplasm) it is possible they interact with the thyroid receptors there. What might happen if they actually do this? 

The thyroid hormone T3 plays an important role in maintaining metabolic balance.  (The different thyroid hormones are closely related; they stimulate growth and metabolism and result in generally increased oxygen consumption and heat production.  This is accomplished by increasing the gene expression at the mRNA protein level of growth hormone and many enzymes including mitochondrial ones). 

The thyroid hormone receptors - which act as transcription factors regulating gene expression - mediate T3 activity. If the thyroid receptors are not bound by thyroid hormones then gene transcription is repressed.  (This suggests that the OASL proteins may be able, by binding with the thyroid receptor, to reduce a wide range of metabolic activities).

The fact that two of the OASL proteins (p56, p59) contain, surprisingly enough, ubiquitin motifs suggests that TRIP 14 could not only bind TR’s but could also target them for destruction.  (Ubiquitin targets a protein for degradation by attaching itself to it. A  proteasome (a protein degrading complex) starts with the strands of ubiquitin and eats anything attached to it.)

A type of communication - called 'cross talking' - where one signaling pathway (IFN) begins at the cell membrane, and ultimately interacts with a totally different pathway (thyroid) in the nucleus, is not common, but does occur. One is present between the TRIP 15 protein and the interferon consensus binding protein (ICSBP) in the nucleus.  TRIP 15 activates two signaling molecules, IxBa and c-Jun, that are important components of the interferon signaling pathway.  TRIP proteins phosphorylate ‘on’ the same serine residues that IBSP does to produce the transcription of genes induced by IFN’s.  TRIP phosphorylation on that specific residue results in repression of the interferon stimulated response elements (apparently stopping IFN transcription in its tracks).  ICSB is expressed only in immune cells and is induced by IFN-y.  Interrupting this pathway results in Th1 deficient mice probably because of their inability to produce IL-12.

(This presents a model that could explain not only the altered response to glucocorticoids evidenced in CFS, but also the low IL-12 production and the deficient Th1 response.  In this scenario, the 2-4OASL proteins which share the binding site but lack the catalytic site of 2-5OAS, (and presumably are unbound just as 2-5OAS is), target the thyroid receptors for destruction.)

Inducing the production of 2-5OASL proteins is complex; it requires the proper balance of IFN subtypes and the activation of PKC (protein kinase C) via PI-3k.  Along with severe fatigue, CFS is characterized by immune disjunction and a hypersensitivity to glucocorticoids that is probably initiated at the gene transcription level.  Given the strong dysregulation of the interferon 2-5A pathway already evidenced in this disease, it is reasonable to consider that a comparable dysregulation in the interferon signaling pathways exists that may be responsible for the resistance to thyroid hormones seen in CFS. 

An inability to respond appropriately to thyroid hormones could result in extreme fatigue even when thyroid hormone levels are normal.  (The authors are suggest that dysregulation in the thyroid receptors in the nucleus of the cell is stopping the activation of the array of genes that the thyroid hormone normally activates)  Upregulation of the IFN 2-5A pathway and a concomitant upregulation of the 2-5OASL proteins/TRIP 14 proteins could possibly result in the destruction of the thyroid receptors and account for the resistance to glucocorticods found in CFS.

 RNase L AND SIGNAL TRANSDUCTION

RNase L’s role in the signaling pathways has begun to emerge recently. RNase L appears to participate in activation of the MAPK and JNK signaling pathways, and RNase L has recently been linked to the regulation of the IFN signaling pathway.  RNase L cleaves both the mRNA’s produced by ISG-15 (an interferon stimulated gene) and by a gene (ISG 43) induced by IFN’s which codes for a protease which interacts with ubiquitin.   Because ISG15 is an immunoregulator, reducing levels of the ISG15 protein will only exacerbate the immune irregularities found in CFS.  Since it is likely that the 37-kDa is regulated differently than 80-kDa RNase L, it is clear, given RNase L’s wide ranging activities, that the dysregulation likely to ensue from the activity of the 37-kDa fragment could have dramatic effects. 

The cleavage of RNase L results in fragments other than the 37-kDa one, one of which contains the N-terminal portions of the protein. A ‘blast search’ indicated that this fragment had a high degree of similarity with three other human proteins, one of which was, interestingly enough, another thyroid receptor interacting protein (TRIP 9).  TRIP 9 contains the motif that enables it to bind with thyroid receptors.  The RNase L fragment contains a very similar motif at a different section of the amino acid sequence. One of these fragments shares ‘up to’ 50% similarity with protein kinase 6 (which blocks cell apoptosis.  Finally the catalytic domain has a high degree of similarity to Ire-1 (chap 2, sec. 2.6). Thus, other fragments produced during RNase L's breakup could negatively effect thyroid functioning as well.  

At the end of this section the authors state that they believe that the possibility that the other fragments generated by the breakup of the native RNase L may interfere with the signaling pathway of the thyroid receptor, (or may interact with membrane receptors via ankyrin repeats), or may inhibit apoptosis, while speculative, merits further investigation.

CONCLUSIONS AND PROSPECTS

Abnormalities in the HPA and IGF signaling pathways may contribute to the pathogenesis of CFS. Three areas in particular may be involved.  A dysregulated 2-5OAS pathway that caused the induction of the 2-5OASL/TRIP  proteins could lead to resistance to thyroid hormones (via the destruction of the thyroid receptor).  Dysregulation of the IFN/2-5A and HPA signaling pathways could result in impaired immunomodulation, cognitive problems and cardiac function. 

(Growth Hormones by and for the Layman.- Growth hormone is essential for protein synthesis and hormone activity and is important in the healing process.  Growth hormone deficiency is typically manifested by increased body fat and waist to hip ratio and decreased lean body mass, extracellular H20 and bone density, and poor concentration, memory, increased irritability and fatigue. 

Reduced extracellular water is attributed to decreased activity of the Na+K ion pump.  GH reduces renin and aldosterone, a steroid hormone that regulates the salt/water balance in the plasma. 

Significant increases in serum triglycerides and decreases in HDL cholesterol (the bad cholesterol) are seen in GH deficiency.  Increased fatigue in GH deficient  adults may be due to altered muscle glucose utilization since IGF stimulates glucose transport into skeletal muscle and stimulate glycogen synthesis. 

The increased energy and expenditure found after GH therapy is partially due to increased conversion of thyroxine, a thyroid hormone, to its metabolite.  Increased feelings of well-being after GH therapy could be due to improved cerebral blood flow , glucose utilization, or the direct effects of GH on the hypothalamus or by IGF on the central nervous system.

A disordered IGF-1 system could effect cell growth, apoptosis, protein synthesis, hormone activity, etc.  IGF disregulation appears to be initiated by low growth hormone levels but could exacerbated in the peripheral circulation by the low DHEA levels found in CFS.  

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