Open AccessCyclin‐dependent kinase inhibitors and basement membrane interact to regulate breast epithelial cell differentiation and acinar morphogenesis
Author(s) -
Coppock H. A.,
Gilham D. E.,
Howell A.,
Clarke R. B.
Publication year - 2007
Publication title -
cell proliferation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.647
H-Index - 74
eISSN - 1365-2184
pISSN - 0960-7722
DOI - 10.1111/j.1365-2184.2007.00463.x
Subject(s) - microbiology and biotechnology , biology , cell growth , basement membrane , cellular differentiation , matrigel , cell , biochemistry , gene
. Objective : The cyclin‐dependent kinase inhibitors (CDKIs), p21 CIP1 and p27 KIP1 regulate growth and differentiation in diverse tissue types. We aimed to determine whether p21 CIP1 or p27 KIP1 could induce a terminally differentiated phenotype in breast cells, and to examine if CDKI expression is regulated by basement membrane interactions. Materials and Methods : Effects of increased CDKI expression on the phenotype of MCF‐10A breast epithelial cells were examined by retroviral transduction of p21 CIP1 or p27 KIP1 cDNA. Results : Overexpression of p21 CIP1 or p27 KIP1 reduced MCF‐10A growth rates in monolayer cultures, altered cellular morphology and stimulated accumulation of neutral lipid droplets, suggesting partial lactational differentiation. However, markers of luminal differentiation (oestrogen and progesterone receptors, α‐lactalbumin, β‐casein and adipophilin) were absent when examined by reverse transcriptase‐polymerase chain reaction and immunohistochemistry. Cell‐basement membrane contacts are known to be essential for full mammary epithelial cell differentiation and therefore parental MCF‐10A cells were cultured on a basement membrane preparation (Matrigel) in which they form acini. Immunocytochemistry showed that Ki67, the cell proliferation marker, was initially expressed at high levels and as growth decreased p27 KIP1 expression steadily increased. Surprisingly, p21 CIP1 was highest at the early stages of acinus growth and was detected in proliferating cells, as demonstrated by colocalization in dual Ki67/p21 CIP1 immunofluorescence. Overexpression of p21 CIP1 or p27 KIP1 impaired formation of acini, whereas their knockdown, using siRNA, increased acinus formation. Conclusion : We conclude that both p21 CIP1 and p27 KIP1 induce partial secretory differentiation of mammary cells in monolayer, but during acinus morphogenesis in 3D culture they have a highly regulated temporal expression pattern.