Proline-Rich Tyrosine Kinase 2 Mediates Gonadotropin-Releasing Hormone Signaling to a Specific Extracellularly Regulated Kinase-Sensitive Transcriptional Locus in the Luteinizing Hormone β-Subunit Gene

Abstract

G protein-coupled receptor regulation of gene transcription primarily occurs through the phosphorylation of transcription factors by MAPKs. This requires transduction of an activating signal via scaffold proteins that can ultimately determine the outcome by binding signaling kinases and adapter proteins with effects on the target transcription factor and locus of activation. By investigating these mechanisms, we have elucidated how pituitary gonadotrope cells decode an input GnRH signal into coherent transcriptional output from the LH beta-subunit gene promoter. We show that GnRH activates c-Src and multiple members of the MAPK family, c-Jun NH2-terminal kinase 1/2, p38MAPK, and ERK1/2. Using dominant-negative point mutations and chemical inhibitors, we identified that calcium-dependent proline-rich tyrosine kinase 2 specifically acts as a scaffold for a focal adhesion/cytoskeleton-dependent complex comprised of c-Src, Grb2, and mSos that translocates an ERK-activating signal to the nucleus. The locus of action of ERK was specifically mapped to early growth response-1 (Egr-1) DNA binding sites within the LH beta-subunit gene proximal promoter, which was also activated by p38MAPK, but not c-Jun NH2-terminal kinase 1/2. Egr-1 was confirmed as the transcription factor target of ERK and p38MAPK by blockade of protein expression, transcriptional activity, and DNA binding. We have identified a novel GnRH-activated proline-rich tyrosine kinase 2-dependent ERK-mediated signal transduction pathway that specifically regulates Egr-1 activation of the LH beta-subunit proximal gene promoter, and thus provide insight into the molecular mechanisms required for differential regulation of gonadotropin gene expression.

Growth Factor Signals in Neural Cells

“…A principal strategy in achieving this selection specificity is compartmentalization of signaling enzymes (43). Another signaling mechanism that we have recently described (45,46) is the creation of discrete signaling microcomplexes linked to transmembrane receptors, such as G protein-coupled receptors, that are able to then translocate to different cell compartments after separating from the cell surface receptor and then effectively transmit a unique signal depending upon the nature of the initial receptor stimulation. One can think of these microcomplexes as encryptons as through their physical constitution and protein stoichiometry these signaling microcomplexes of common proteins assemble in a manner that allows functional encryption of the exact nature of the initial input ligands (45).…”

Conservation of mechanisms mediating gonadotrophin-releasing hormone 1 stimulation of human luteinizing hormone β subunit transcription

“…Although both the ERK1/2 and JNK MAPK cascades have been implicated in GNRH1 regulation of the Lhb promoter in other species (Yokoi et al, 2000;Harris et al, 2002), a more critical role has been attributed to ERK1/2 (Liu et al, 2002;Kanasaki et al, 2005). GNRH1 stimulates Egr1 expression through the ERK1/2 pathway (Duan et al, 2002;Maudsley et al, 2007), and EGR1 appears to be the primary transducer of the GNRH1 signal to the Lhb promoter (Dorn et al, 1999;. Indeed, our data confirm a critical role for EGR1 in regulation of the human LHB promoter through two conserved cis-elements.…”

Extracellular Matrix Proteins in the Anterior Pituitary Gland

“…The ECM has been noted to be important in activation and transmission of signals in gonadotrophs [79]. In particular, actions of GnRH via GnRH-receptor that result in integrin regulation have been detected, whereby GnRHactivated pathways have been demonstrated to induce temporal changes in the composition of the signalling complex that seems to reside in microtubules, and directs the signalling complex along the cytoskeleton to the network of focal adhesions [80] to mediate cell adhesion.…”

“…The focal adhesions, which alter cytoskeleton and LH-transcription [81], can be disrupted by cytoskeletal inhibitors. Some details have been delineated, and the involvement of Pyk2 tyrosine phosphorylation with ERK activation and c-Src are implicated in the network of cytoskeleton-mediated processes [82]; these steps themselves being regulated by binding of GnRH to the GnRH-R (see [79]). Hence structural integrity of the cell cytoskeleton is central to cell function including GnRH-mediated cell migration and spreading.…”