RNAi of FACE1 protease results in growth inhibition of human cells expressing lamin A: implications for Hutchinson-Gilford progeria syndrome

Authors:
Gruber J, Lampe T, Osborn M and Weber K
In:
Source: J Cell Sci
Publication Date: (2005)
Issue: 118(Pt 4): 689-696
Research Area:
Cancer Research/Cell Biology
Cells used in publication:
HL-60
Species: human
Tissue Origin: blood
Platform:
Nucleofector® I/II/2b
Abstract
FACE 1 is the endoprotease responsible for cleavage of prelamin A to lamin A. Transfection of HeLa cells with siRNA for human FACE 1 results in a strong phenotype. Protein and mRNA levels for FACE 1 are knocked down and cell division stops abruptly. Two populations of cells are detected. The first form aberrant mitotic spindles, arrest in mitosis and later enter apoptosis. The second show dramatic changes in nuclear morphology with extensive formation of lobulated nuclei and micronuclei. Using antibodies that specifically recognise prelamin A, but not lamin A, we show that prelamin A accumulates at the nuclear lamina in FACE1 silenced cells, whereas in control cells prelamin A is found in many small nuclear dots, but not at the nuclear lamina. In double knockdown experiments with FACE 1 and lamin A siRNAs, the results depend on which protein is knocked down first. FACE1 knockdown 24 hours prior to lamin A knockdown gives results similar to the single FACE1 knockdown. By contrast, lamin A knockdown 24 hours prior to FACE1 knockdown results in none of the changes described above. Silencing of FACE1 in HL60, a cell line that lacks lamin A, also has no effect. The combined results suggest that prelamin A is a poison in cells subjected to FACE 1 knockdown. Finally, we draw attention to similarities in phenotype between FACE1-silenced HeLa cells and fibroblasts from patients with Hutchinson-Gilford progeria syndrome containing prelamin A mutations that prevent cleavage by the FACE1 endoprotease.