IRF6 in development and disease: a mediator of quiescence and differentiation

Abstract

Post utero development of the mammary gland is a complex developmental process characterized by states of rapid cell proliferation (branching morphogenesis) followed by functional differentiation (lactation) and the consequent apoptosis (involution) of the secretory mammary epithelial cell. This process is cyclical, such that involution returns the mammary gland to a near-virgin-like state capable of responding to morphogenic cues with each consecutive pregnancy. Importantly, many of the regulatory processes which oversee mammary gland development are corrupted or otherwise compromised during the development of breast cancer. For example, Interferon Regulatory Factor 6 (IRF6) is a novel protein with growth inhibitory properties that was initially identified in mammary epithelial cells through its interaction with maspin, a known tumor suppressor in normal breast tissue. Recent findings from our laboratory suggest that IRF6 functions synergistically with maspin to regulate mammary epithelial cell differentiation by acting on the cell cycle. This perspective focuses on the possible involvement of IRF6 in promoting differentiation by regulating exit from the cell cycle and entry into the G(0) phase of cellular quiescence, and how these new findings shed light on normal mammary gland development and the initiation and progression of breast cancer.

Figure 1. IRF6 phosphorylation is unaffected by the cdk inhibitor flavopiridol

MCF-10A mammary epithelial cells were serum starved for 48 hours. Serum-starved cells were pre-treated with flavopiridol for one hour at a 400 nanomolar concentration prior to the addition of serum. Cells were harvested at 15 and 30 minutes following serum addition according to standard techniques. Phosphorylated and non-phosphorylated forms of IRF6 are demarcated by the arrows. Maspin levels were unchanged and used as a loading control.

Figure 2. IRF6 responds to treatment with the TLR3 agonist polyI:C

A) TLR3 mRNA is detectable by RT-PCR in MCF-10A cells whereas TLR4 mRNA was undetectable. Expression of TLR3 mRNA was reduced in log-phase proliferating cells (growth) and increased in confluent cells. Treatment of MCF-10A cells with the TLR3 agonist polyI:C (a synthetic double-stranded RNA construct) induced an increase in TLR3 mRNA whereas lipopolysaccharide (LPS), an agonist for TLR4, did not affect TLR3 mRNA expression. B) Western blot analysis of MCF-10A cytosolic and nuclear fractions following treatment with polyI:C. IRF6 is maximally expressed in the nuclear fraction 30 minutes after polyI:C treatment. Keratin 18 was used as a cytosolic marker and Lamin b1 was used as a nuclear marker. Fractionation was performed as previously described.

Figure 3. A model illustrating the involvement of IRF6 in promoting quiescence and differentiation by instigating entry into the G(0) phase of cellular quiescence

Increasing IRF6 levels promote exit from the cell cycle. As cells proliferate, IRF6 is ubiquitinated and targeted for proteasomal degradation.