Clinical Biochemistry of Domestic Animals (Sixth Edition) - UMK ...

568
Chapter | 18 Pituitary Function
Pig SYSMEHFRWGKPVGKKRRPVKVYPNGAEDELAEAFPLEF 39
Dog ------------------------------S-----V--
Cattle ------------------------------S-Q------
Sheep ------------------------------S-Q------
Human ------------------------------S--------
Rat -------------------------V--N-S--------
Mouse -------------------------V--N-S--------
************************* ** * * *** **
Cattle FIGURE 18-6 Comparison of the ACTH
sequences from various species using the
Sheep
one-letter code for amino acids. Identical
Dog amino acids are given by a single line (-).
Pig
The asterisk (*) means an identical sequence
for all species. The shaded box indicates the
Human two pairs of basic amino acids prone to proteolytic
cleavage. The insert shows a dendrogram
indicating the mutual Mouse
relationship.
Rat
1 . ACTH/ α -MSH
a . Gene Expression
The gene encoding POMC contains three exon areas.
Exon 1 encodes a 5 untranslated region (5 -UTR), and
exon 2 encodes the signal peptide and the N-terminal part
of the POMC prohormone. The majority of the prohormone,
including γ -MSH, is encoded by exon 3. After the splicing
process, an mRNA of approximately 1150 nucleotides
is formed in the pituitary of most species, including the
dog ( Mol et al. , 1991 ). In many peripheral tissues, very low
expression of shorter POMC mRNA sequences are found,
whereas in nonpituitary tumors, an mRNA of 1350 nucleotides
can be found ( Newell-Price, 2003 ).
The presence of an unmethylated CpG island in the
POMC promoter is a prerequisite for gene expression,
whereas methylation completely blocks gene expression
( Newell-Price, 2003 ). These methylation patterns are set
during early development. Synergistic stimulation of gene
expression requires binding of yet partially characterized
transcription factors in the distal and central part of the
POMC promoter. The expression of the POMC gene in corticotropes
and melanotropes is regulated by the same promoter
elements as shown by experiments with transgenic
mice ( Tremblay et al. , 1988 ). Three important response elements
act within the central promoter region. The pituitary
homeobox 1 (Ptx1) protein, which is widely expressed in the
pituitary, acts in synergy with the corticotrope/melanotropespecific
factors NeuroD1 ( Newell-Price, 2003 ) or Tpit
(Quentien et al. , 2006 ) to stimulate POMC gene expression.
Binding of corticotropin-releasing hormone (CRH) stimulates
POMC expression in the PD through stimulation of
cAMP within the cell ( Jacobson and Drouin, 1994 ). CRH
acts synergistically with the leukemia-inhibitory factor (LIF)
through binding of the cAMP response element binding protein
(CREB) at the response element sites for NUR77 and
STAT1-3 ( Mynard et al. , 2004 ). Nur77 is an orphan nuclear
receptor that antagonizes the negative feedback of glucocorticoids
on POMC expression ( Martens et al. , 2005 ).
POMC expression in the IL melanotrope is also stimulated
by cAMP. In vitro experiments in various species point
to a possibility that CRH activates this signal transduction
system. However, chronic infusion of CRH stimulates the
POMC mRNA concentrations in the AL but inhibits mRNA
concentration in the IL ( Hollt and Haarmann, 1984 ). IL
POMC expression is also regulated by stimulation with serotonin
and norepinephrine or GABAergic and dopaminergic
inhibition ( Saland, 2001 ). Glucocorticoids inhibit AL POMC
expression but have no effect on the IL expression of POMC.
Extrapituitary POMC expression is found in many tissues
( DeBold et al. , 1988 ), also in skin tissue of various
species including the dog ( Kidney et al. , 2004 ).
b . (Pro)hormone
POMC prohormone sequences are known for a variety of
species and share a common structure ( Fig. 18-5 ). Among
species a high sequence homology is found between the
N-terminal site (N-POC) of the prohormone (including
γ -MSH), the ACTH region, and the β -endorphin region.
In contrast, the regions between γ -MSH and ACTH and
the first part of β -lipotropin are rather heterogeneous ( Mol
et al. , 1991 ; Numa and Imura, 1985 ). Species differences
in the sequence of ACTH occur only in the N-terminal part
( Fig. 18-6 ), whereas the ACTH(1-24) sequence, necessary
for full biological activity, is identical among mammals.
The specificity of immunoassays to measure ACTH
concentrations is achieved by directing antibodies toward
the ACTH (13-18) epitope, which is absent in MSH and
corticotropin-like intermediate lobe peptide (CLIP). These
antibodies do, however, cross-react with intact POMC
and may result in apparent elevated ACTH concentrations
( Goossens et al. , 1995 ). Two-sided assays are more sensitive
to variation in the carboxy-terminal part and show
insufficient cross-reactivity among the species.
The proteolytic enzymes PC1/PC3 and PC2 are
involved in the processing of prohormones in neuroendocrine
cells. They cleave the precursor at pairs of basic
amino acids, resulting in the formation of biologically
active hormones. The PC1/PC3 enzyme present in the AL
cleaves POMC to ACTH and β -LPH. PC2 generates the
ACTH(1-13) fragment in both the AL and IL. The combination
of both enzymes is present in the melanotropes of
the IL. In the hypothalamus, these enzymes determine also
the availability of POMC-derived melanocortins, which
play an essential role in the regulation of energy balance
( Bertile and Raclot, 2006 ; Helwig et al. , 2006 ).
In the melanotrope α -MSH is formed by C-terminal
amidation and N-terminal acetylation of ACTH(1-13).
Various degrees of acetylation of MSH result in the storage