New reports indicate a chromosomal rather than hormonal basis for the susceptibility of females to autoimmune disease. It is held that if females reactivate an inactivated X chromosome there will be overexpression of certain X-located genes affecting immune function. Hence, normal mechanisms of self/not-self discrimination might be impaired resulting in immune reaction to self antigens. 

    However, the data are also consistent with the long-held view that the demands of intracellular self/not-self discrimination have driven the evolution of X-chromosome dosage compensation. It was proposed that, whether cells are in male or female bodies, concentrations of proteins are fine-tuned up to their aggregation thresholds. A disruption of this equilibrium, by agents originating either externally (e.g. virus) or internally (e.g. reactivated X chromosome), generates homoaggregates that trigger responses against the respective not-self or self antigens. 

   Thus female susceptibility to autoimmune disease may not be because certain immune system genes happen to be X-located, but because self/not-self discrimination was the raison d'etre for X chromosome dosage compensation in the first place.

Keywords: autoimmunity; dosage compensation; double-stranded RNA; protein aggregation; systemic lupus erythematosus; X chromosome

I have proposed that a need for intracellular self/not-self discrimination drove the evolution of X chromosome dosage compensation.^1,2 New reports, implicating sex chromosomes rather than hormones in the female predisposition to autoimmune disease,^3-6 suggest an update will be helpful.

 In human females one X chromosome is inactivated so that, like males, each cell has only one functional X chromosome. This phenomenon, seen in various forms across a wide range of species, equalizes the concentrations of X chromosome gene products (for present purposes proteins) between males and females, and is referred to as "dosage compensation." The value-laden term "compensation" implies that intolerance to dosage differences has been beneficial, thus ensuring the differential survival of organisms that evolved mechanisms to bring it about. However, why compensation should be adaptively advantageous - a paradox noted by Muller⁷ - receives little attention.

    One possibility is that a concentration-dependent function to which many proteins contribute is likely to be favored by dosage compensation. Thus a given protein has both a specific function (e.g. enzymic) and a collective function (e.g. contributing to total protein concentration). The concentration of the protein should be sufficient to meet both specific and collective needs. An example of a collective function is the Gibbs-Donnan equilibrium by which intracellular proteins influence the distribution of ions across cell membranes.^8,9 Failure precisely to regulate protein concentrations could perturb cell membrane potential which, in turn, could impair various processes that depended on that potential. Thus, over evolutionary time rates of protein synthesis and/or degradation within cells would have been fine-tuned to sustain constant protein concentrations from generation to generation, whether the cells are in male or female bodies. We are here concerned with another, largely unrecognized, collective protein function, which relates dosage compensation to self/not-self discrimination and hence to immune function.^1,2

 Invading viruses and bacteria normally mobilize cells of the immune system (lymphocytes, phagocytes) and the resulting inflammatory response (often seen as reddening and swelling) acts to limit the spread of, and destroy, the foreign organisms. The term "autoimmune" is applied to diseases that are considered to result from a failure to distinguish immunologically "self" from "not-self" (foreign). In systemic lupus erythematosus (SLE) many tissues become inflamed without the obvious involvement of a foreign organism. Complement components promote the inflammation, but they may also be important in deleting the self-reactive lymphocytes that initiate the process.¹⁰ Thus, the incidence of SLE is higher when there is complement-deficiency.^11,12

    The fact that the incidence of SLE in human males is nine-fold less than in females, where the onset is usually post-pubertal and prior to menopause, has suggested an influence of female hormones.¹³ However, Klinefelter syndrome males (chromosomally XXY) are also predisposed to SLE, implying a direct X chromosome gene dosage effect.³ Furthermore, studies in mice, where manipulation of the testis-determining Sry gene has permitted separation of chromosomal and hormonal backgrounds, have implicated the X chromosomes themselves in the female bias towards experimental autoimmune encephalitis and SLE.⁴ Consistent with this, SLE has a low incidence in Turner's syndrome (XO).

Such considerations have given rise to the hypothesis of interference with normal dosage compensation by reactivation of the inactive X chromosome in certain females and certain Klinefelter males.^5,6 This is in keeping with reports that the DNA methylation necessary for the inactivation is defective in SLE.¹⁴ Thus, doubling the activities of certain X-linked genes whose products are involved in immune processes might explain the susceptibility to SLE.¹⁵ However, the defect in DNA methylation should have a genome-wide impact. Accordingly, without a correlated increase in cell volume, the cytosolic concentrations of many proteins, especially those encoded by the X chromosomes, should increase. While the specific activities of certain proteins would be increased, there should also be an influence on collective protein functions - perhaps on one concerned with intracellular self/not-self discrimination.

 A possible mechanism for intracellular self/not-self discrimination implicating a collective protein function was developed from four premises:

(i) Single celled organisms arose before multi-cellular organisms.

(ii) There existed pathogens capable of gaining entry into single celled organisms.

(iii) The cells developed internal systems for distinguishing pathogen molecules from their own.

(iv) Multi-cellular organisms adapted these pre-existing systems when evolving their own immune defences.

The case, elaborated elsewhere,^16-22 will be briefly summarized.

    As it approaches the limits of its solubility (i.e. if its concentration is progressively increased) a given macromolecular species (protein) will form, with high specificity, relatively insoluble like-with-like homoaggregates.^23,24 In the crowded cytosol, with a total protein concentration around 300mg/ml,²⁵ the concentration of a protein affects not only its own solubility but also the solubilities of other proteins. Thus, a group of proteins exerts a collective "pressure" tending to make individual protein species self-aggregate. A protein species with the greatest propensity to self-aggregate (a function of factors such as structure, molecular size, and initial concentration) will aggregate first. As the pressure (i.e. total protein concentration) increases, other protein species follow suite. Each individual protein species both contributes to, and is influenced by, this pressure. Each protein has this collective function as well as its own specific function. Both functions affect phenotype and hence influence selection by evolutionary forces.²⁶

    So, over evolutionary time, the concentration of each protein has been fine-tuned to the concentrations of the other proteins with which, from generation to generation (be it male or female) it has shared a common cytosol. Sex chromosome dosage compensation is part of this process. Each protein, through its personal degree of solubilization, pleiotropically influences the solubilization, and hence activities, of numerous other proteins with which it may not otherwise be functionally related.²⁷ The homeostatic fine-tuning of the concentration of a protein to that of its companion proteins implies high conservation of the degree of macromolecular crowding.²⁸

    When a foreign pathogen (e.g. virus) enters a cell this equilibrium is perturbed. Whereas self protein-encoding genes are at peace with the concentrations at which their products have arrived, the raison d'etre of most viruses is, at some time within the lifespan of their host, to increase in number. This implies the synthesis of virus-specific proteins at rates that may result in unacceptably high cytosolic concentrations. This would cause aggregation of virus proteins (supplemented by coaggregation of some host proteins), to a degree that could constitute a not-self signal. This, in turn, would trigger various adaptive options, one of which would be apoptosis.

    However, given that most viruses have higher mutation rates than their hosts, viruses would soon adjust their synthesis rates to avoid triggering host defence mechanisms. Thus, the aggregation-triggered defence mechanism should mainly serve to limit the rate of virus growth, so slowing its multiplication and spread to other host cells. Supporting roles of molecular chaperones, heat-shock proteins and nucleic acids in this process have been proposed.^16-22 Recently, the emergence of a "hidden transcriptome" from what was formerly regarded as "junk DNA",^29-31 has lent credence to the hypothesis of an independent role for nucleic acids in self/not-self discrimination.^19,32,33

Double-stranded RNA has long been known as an alarm signal that activates immune defenses.^32-35 Indeed, viruses often first express themselves in foreign cytosols in the form of RNA transcripts that can fold into stem-loop structures. If double-stranded stem segments of a particular length, which originated entirely from a virus, were sufficient to alert a host, then it is likely that viruses which had mutated to prevent duplexes of that length from appearing or being detected,^32,33,36 would have preferentially survived. Hence, on a priori grounds, duplex formation between host RNA and virus RNA seems more plausible. It was proposed that the spectrum of host cytosolic RNAs should be regarded, not in terms of their individual specific functions (e.g. as mRNAs specifying proteins), but as an RNA collective that is best equated with the antibodies which mediate extracellular immune responses in higher organisms.^19,22 However, whereas antibodies (immunoglobulins) are dedicated, each to a range of related antigenic determinants with varying affinity, many intracellular RNA "antibodies" have additional roles, such as encoding protein. When there is a crisis - invasion by a virus - the latter roles would become of relatively minor importance (see later).

   Thus, host transcripts can be regarded as presenting a wide range of specificities for RNA targets. Some host transcripts have the potential to recognize, and pair to form lengths of double-stranded RNA with, sequences in the genomes or transcripts of viruses. The recognition process involves initial "kissing" interactions between RNA secondary structures, which is followed by duplex formation in the regions of complementarity.^19,22 Furthermore, just as self/not-self discrimination at the protein level might be subverted by inadvertent expression of autologous proteins due to failed X inactivation (see above), so self/not-self discrimination at the RNA level might also be subverted by inadvertent formation of self duplexes, with pathologically "toxic" results.^37,38

   While the evidence for the RNA "antibody" role in eukaryotes is at present fragmentary,^19,22,33 there is now compelling evidence in prokaryotes. To deal with bacteriophages, bacteria have long been known to distinguish self from not-self at the DNA level (i.e. restriction enzymes). A distinction is also made at the RNA level by an RNA antibody-like mechanism.^39,40 By mutating its target sequences a phage can evade this bacterial defense system. Since secondary structure is required for the initial recognition event, mutations near the target sequence that interfere with phage secondary structure should also be evasive. Host defenses can indeed be evaded by mutating the non-targeted flanking sequences.⁴¹

   In a multicellular organism it can be supposed that, following a primary self/not-self discrimination event at the RNA level, host "alarms would ring" (e.g. interferon induction and upregulation of MHC protein synthesis). Certain heat-shock proteins would shift from chaperone-mode (i.e. disaggregation mode), to aggregation-mode.^17,42 The above aggregates of foreign or self proteins would then be directed to proteosomes,⁴³ leading to their preferential degradation to peptides that would associate with MHC proteins and be displayed at the cell surface for recognition by cytotoxic T cells.²²

The magnitude of the aggregation in SLE is such that there is often systemic involvement of many tissues. However, in certain diseases aggregation is restricted to a limited number of gene products.⁴⁴ Here there might be an opportunity for multiplication of neighboring cells so that a cell targeted for T-cell destruction would be quickly replaced. Overt inflammation might not be apparent and cells with visible aggregates (inclusion bodies) might not be detected. However, T cells have limited access to the central nervous system, which is less able to replace cells that are destroyed. Thus, although these diseases may be systemic, they could appear to target the nervous system specifically.³²

A self/not-self alarm ("stress") invokes collective RNA and protein functions that should normally operate independently of any specific roles of individual RNA and protein species (see above). Thus, collective functions trump specific functions. However, as part of collective responses specific proteins could either increase (following increased general transcription) or decrease (following coaggregation) to extents sufficient to influence phenotypes. For example, malexpression at the wrong time or place of a key regulatory protein (e.g. during embryonic development) could produce unwelcome "phenocopies" of forms normally encountered as inheritable mutants.^20,45

The proposal that the aggregation mechanism for self/not-self discrimination initially evolved at the unicellular level and was subsequently adapted by multicellular organisms, implies a fundamental process that should apply universally whenever an aneuploid threat to dosage balance arises.^2,46 Yet, sex chromosome dosage compensation in birds has long been in doubt.^47,48 However, recent studies suggest that birds dosage-compensate on a gene-by-gene basis - locally rather than globally.⁴⁹ Indeed, even in organisms where compensation mechanisms appears global, certain genes are compensated more than others.⁵⁰ The present hypothesis predicts that genes whose products most powerfully exert aggregation pressure (e.g. large size and high abundance) are most likely to be compensated than those less powerful in this respect (e.g. small size and low abundance).⁵¹ Since aggregation propensity increases with temperature,²³ pyrexia in the early stages of an infection should be beneficial. By the same token, the severity of autoimmune disease should be decreased by vigilant employment of antipyretic medication.

Hypotheses on the hormonal basis, and on the mono- or oligo-genic specific immune basis, of the female predisposition to autoimmune disease, now appear less persuasive than the polygenic hypothesis. Under the latter, independently of their specific functions, proteins collectively exert an aggregation pressure the homeostatic regulation of which is perturbed, either by foreign agents (normal immune response), or by failure to dosage compensate an X chromosome (autoimmune response). This intracellular system for self/not-self discrimination works in concert with a double-stranded RNA-based discrimination system. A possible role for other collective macromolecular functions (e.g. Gibbs-Donnan equilibrium) has not been excluded.

Queen's University hosts my web-pages where some of the cited articles may be found.  

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3. Scofield RH. Bruner GR. Namjou B. Kimberly RP. Ramsey-Goldman R. Petri M. et al. Klinefelter's syndrome (47, XXY) in male systemic lupus erythematosus patients. Support for the notion of a gene-dose effect from the X chromosome. Arth Rheum 2008; 58: 2511-2517.

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"This paper is very badly flawed by selective referencing of the literature and failure to think about how its hypothesis applies beyond SLE. Thousands of papers are available on the genetics of SLE and on the genetics of other autoimmune diseases that link them to chromosomes other than the X chromosome. Thousands of papers have been written on animal models of SLE and other autoimmune diseases that initiate autoimmune diseases without the viral or retroviral invasion that the author's hypothesis requires to initiate protein aggregation. Thousands of papers have looked at susceptibility to autoimmune diseases in humans that do not show the huge sex difference in susceptibility to autoimmunity that SLE does. So at a minimum, the author MUST address the vast amount of data that do NOT fit his hypothesis: why do millions of xy males develop autoimmunity? why are viral infections unnecessary for initiating most animal models of autoimmunity? why are there so many non-x-chromosome genes associated with autoimmunity risk? etc. Theories don't just gather supporting evidence: they must also address apparently contrary evidence -- and in this case almost all the evidence fails to fit!"