Discuss dioxins is via food, especially meat

Discuss both the structure of the aryl hydrocarbon
receptor and its role in chloracne

Dioxins,
the most potent of which is 2,3,7,8-tetrachlorodibenzo- p -dioxin (TCDD), form
a group of persistent organic pollutants present in the environment. They
accumulate in the food chain, especially in the animal fat, and more than 90%
of human exposure to dioxins is via food, especially meat and dairy products. The
aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor
found in the cytosol/nucleus and controls the expression of a diverse set of
genes. TCDD is almost exclusively mediated through this receptor. However, the
key alterations in gene expression that mediate toxicity are poorly understood.
This essay will first discuss the structure of AhR receptor and then explain
its role in chloracne.

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Structure of AhR

The
AhR and the AhR nuclear translocator (ARNT) belong to the family of basic helix-loop-helix-PER-ARNT-SIM
(bhlh- PAS) domain containing transcription factors. ARNT shares homology with drosophila
proteins PER and SIM. The PAS-ARNT-SIM domain contains PAS-A and PAS-B regions
of ~130 amino acids each, further, the ligand-binding region is in the PAS-B
domain (Wu et al. 2013). ARNT 1 is ubiquitously present whereas ARNT is
expressed in the brain and kidney.

The
Bhlh component is a well-characterised DNA binding protein motif composed of
two a-helices connected by an intrinsically unstructured loop. They have the
ability to heterodimerise with the members of class I bHLH-PAS group e.g.
hypoxia inducible factors (Hif-1a/ Hif-2a) and the single-minded proteins
(SIM1/SIM2) to regulate a transcriptional response to xenobiotic.

On
the other hand, AhR is a cytosolic signal sensor, which is guided into the
nucleus upon binding of planar aromatic hydrocarbons e.g. TCDD. Further, TCDD
mediates its toxicity through AhR activation (Poland et al. 1976). A
transactivation domain (TAD) involved in the recruitment of other
transcriptional co-activators.

Structurally
the AhR and ARNT are very similar as demonstrated by the diagrams below:

 

bHLH
dimerization dna binding

                            PAS secondary dimerization                                                                                              Transactivation

 

 

AhR                                                                                                           Xenobiotic response

 

                       

15aa

 
 

 

 

 

 

ARNT 1&2                                                                     
                    General partner factor

TCDD

To
date, the pathogenesis of Chloracne remains incompletely understood. However,
most of the effects of TCDD in humans and animal models seems to be mediated by
the AhR signalling pathway.

It
is a planar halogenated aromatic hydrocarbon with high affinity for AhR;
furthermore, it is highly lipophilic and shows low basal rates metabolically. Chloracne
is both human and TCDD specific, it contains acne vulgaris like lesions
including open and closed comedones and cysts, and however unlike acne vulgaris
the lesions are sterile and non-inflammatory. Chloracne consists of
abnormalities in keratinocyte differentiation in the follicle, causing thickened
and compacted stratum corneum, which leads to follicular plugging and
characteristic comedones. AhR was previously found to be expressed and
activated in human keratinocytes and skin fibroblasts as well as in human
sebaceous glands. In the skin lesions of chloracne patients exposed to dioxin,
there was an activation and upregulation of p- epidermal growth factor receptor
EGFR (EGFR), pmitogen-activated protein kinase, and CK17 in all chloracne
tissues but not in the controls. Furthermore, an increase in the dioxin
independent transcriptional activity of the AhR has recently been demonstrated
in a drosophila model to be also responsible for the in vivo dioxin toxicity
(Ju et al,. 2011).

Furthermore,
Acne affects young adolescents whereas chloracne can affect any age group,
anatomical localisation in the former is in the face, but the latter has a
wider variation of targets including the ears and the groin for example. In
acne, sebum production is increased, inflammation is common, limited comedones
and the sebaceous gland is hypertrophic. Conversely, in chloracne the sebum
production is decreased, inflammation is rare, increased blackheads, and
atrophy of the sebaceous gland is seen.

There
are animal models that exhibit chloracne-like disease (for example, the
hairless mouse model (Panteleyev et al., 1997), but because of differences in
the hair follicle (the main site of TCDD induced changes causing chloracne)
between human and animal, extrapolation from the animal models to human disease
is difficult. Chloracne is characterised by hyperplasia of the epidermis,
keratinisation of the pilsebaceous unit, hyperplasia/metaplasia of the
sebaceous gland.

In
its activated state the AhR resides in the cytoplasm in a complex with its
chaperone proteins; the AhR interacting protein (AIP) and heat-shock protein
(hsp90). This complex has multiple functions involving keeping the AhR
localised in the cytoplasm and repressing conformational changes that occur
during activation. AIP/XAP2 is an immunophillin like protein that is part of
the AhR-hsp90 complex binding to both AhR and hsp90 in the absence of ligand
(Bell and Poland, 2000). AIP binding to the AhR inhibits nuclear translocation
by inhibiting importin ? recognition of the NLS (Petrulis et al., 2003) and
ligand binding therefore breaks the XAP2-hsp90-AhR complex down which is
thought to be required for AhR-hsp90 stability, ultimately the conformational
change exposes the N-terminal of NLS, thus initiating nuclear import.

In
the nucleus, hsp90 is exchanged for partner ARNT; which dimerises with the AhR
through aforementioned Bhlh and PAS domains to form a functional DNA binding
complex. Although the stage at which XAP2 and p23 are displaced is vague.  

Yeast
hybridisation and biochemical analysis have been used to demonstrate AhR
interaction with AIP2. Further, transient transfection studies demonstrate a
~two-fold increase in XRE driven reporter gene activity upon AIP2 co-expression
with the AhR, correlating with a ~two- fold increase in AhR protein levels. To
this end, AIP2 decreases ubiquitinated forms of the AhR in transient
overexpression experiments. Treatment of AhR/chaperone complex in-vitro with
geldanamycin destabilises this complex such that p23 and XAP2 are lost from the
complex. Thus supporting the idea that XAP2 is essential for the stability of
the AhR/hsp90/p23 complex, preventing transient unmasking of the N-terminal
nuclear localisation sequence within the AhR diminishing ligand independent
nuclear accumulation. Additionally, the presence of XAP2 in the latent chaperone
complex protects the AhR from proteasome-mediated degradation, an observation
indicative of a stable AhR–chaperone complex.

Since
XAP2 interacts with both hsp90 and the AhR. it is likely that XAP2 acts as a
stabilising factor and the presence of XAP2 in this complex may denote a subset
of mature AhR–chaperone complexes. Recent evidence from Hepa-1 cells confirms
that XAP2 functions to prevent nucleocytoplasmic shuttling, but that its
depletion has no effect on ligand induced signalling in these cells (Pollenz et
al. 2006).  

Downstream
molecular targets of AhR signalling are versatile for example the drug
metabolising enzymes e.g. CYP1A1 and CYP1A2, IL-Ib, Tgf’s, C-myc, EGFRs and
Blimp1. Among the aforementioned targets of AhR, c-Myc and Blimp-1 supposedly
play an important role in modulating directional differentiation of epidermal
progenitor cells and be a potential important target linkage between specific
TCDD-induced skin pathology and the stem-cell differentiation changes in the
skin.

Activation
of c-Myc favours differentiation along the lineages of the interfollicular
epidermis and sebaceous gland, and it results in the appearance of groups of
differentiated sebocytes within the interfollicular epidermis.

Inactivation
of c-Myc results in hypoplasia of the sebaceous glands. Loss of Blimp1 results
in the elevation of c-Myc expression and hyperplasia of the sebaceous glands.45
Taken together, TCCD/AhR/ARNT/Blimp1 and CYP1A1/c-Myc pathways may very likely
be the proposed signalling pathways of TCDD on skin stem cell progenitors, leading
to TCDD-induced abnormal differentiation of sebaceous gland.

 

Treatment and conclusion

Knowledge
and application of treatments are limited since the direct/combined mechanisms
of TCDD toxicity remains unknown and tried methods have proved unsuccessful
(Yip et al. 1993) although perhaps in the future, manipulation of AhR
signalling could be employed. Current options include self-resolution over time
and surgical removal of the comedones. 
Recently, a synthetic dietary fat substitute ‘Olestra’ has shown to have
the ability to bind to chloracnegens and accelerate their faecal excretion and
indeed Geusau et al (1999) have showed that combining olestra and caloric
restriction causes a 30-fold increase in in rate of excretion of a labelled
compound, this may be a prospective therapeutic method in the future.

In
conclusion, much progress has been made regarding the biochemistry and
functional role of AhR signalling in relation to dioxin toxicity however, a firm
mechanism is yet to be elucidated, conversely, there may not be one single
pathological mechanism and rather AhR signalling is a multi-model system that
works together to induce chloracne.