Heterozygous mutations in the PALB2 hereditary breast cancer predisposition gene impact on the three‐dimensional nuclear organization of patient‐derived cell lines

Abstract

PALB2/FANCN is a BRCA1- and BRCA2-interacting Fanconi Anemia (FA) protein crucial for key BRCA2 genome caretaker functions. Heterozygous germline mutations in PALB2 predispose to breast cancer and biallelic mutations cause FA. FA proteins play a critical role in the telomere maintenance pathway, with telomeric shortening observed in FA cells. Less is known about telomere maintenance in the heterozygous state. Here, we investigate the roles of PALB2 heterozygous mutations in genomic instability, an important carcinogenesis precursor. Patient-derived lymphoblastoid (LCL) and fibroblast (FCL) cell lines with monoallelic truncating PALB2 mutations were investigated using a combination of molecular imaging techniques including centromeric FISH, telomeric Q-FISH and spectral karyotyping (SKY). Mitomycin C and Cisplatin sensitivity was assayed via cellular metabolism of WST-1. The PALB2 c.229delT FCL showed increases in telomere counts associated with increased mean intensity compared with two wild-type FCLs generated from first-degree relatives (P =1.04E-10 and P =9.68E-15) and it showed evidence of chromosomal rearrangements. Significant differences in centromere distribution were observed in one of three PALB2 heterozygous FCLs analyzed when compared with PALB2 wild-type, BRCA1 and BRCA2 heterozygous FCLs. No significant consistently increased sensitivity to Mitomycin C or Cisplatin was observed in LCLs. Our results are suggestive of an altered centromere distribution profile and a telomere instability phenotype. Together, these may indicate critical nuclear organization defects associated with the predisposition to transformation and early stage development of PALB2-related cancers.

Distinct Nuclear Organization of Telomeres and Centromeres in Monoclonal Gammopathy of Undetermined Significance and Multiple Myeloma

“…Sarkar et al [25] previously established that, in mice, centromeres are located more toward the nuclear periphery in normal lymphocytes but are more concentrated in the middle of the nucleus in immortalized PreB lymphocytes and even more so in the murine plasmacytoma line MOPC460D [25]. Wark et al [20] showed that the spatial position of centromeres in interphase nuclei was altered in fibroblast cell lines with monoallelic truncating PALB2 mutations (PALB2 c.3323delA) compared to wild-type controls and fibroblasts with BRCA1 and BRCA2 mutations. This difference was found to be a contributing factor in increasing the propensity for chromosomal rearrangements [20].…”

“…The evolutionary hotspots in the pericentromeric chromosomal regions that have been implicated as major contributors to genomic instability and large-scale genetic changes such as chromosomal rearrangements, deletions, and insertions or bursts of retrotransposon activity could force centromere positions to change [16]. However, little is known about effects of those changes in the dynamics and stability of centromeres in cancer [18,19,20]. Consequently, the positioning of the centromeres in the interphase nucleus has been the focal point of several studies (e.g., [19,20,21,22,23,24]).…”

“…However, little is known about effects of those changes in the dynamics and stability of centromeres in cancer [18,19,20]. Consequently, the positioning of the centromeres in the interphase nucleus has been the focal point of several studies (e.g., [19,20,21,22,23,24]). For instance, it has been demonstrated that the distribution of the centromeres in the nucleus of the same cells is altered in the process of immortalization and malignant transformation in both mice and humans [24,25].…”

Interaction with PALB2 Is Essential for Maintenance of Genomic Integrity by BRCA2

“…Recent studies have suggested that heterozygosity of BRCA2 as well as PALB2 results in some loss of DNA replication fork protection and a defect in DNA damage response, which contributes to genomic instability [ 38 , 39 ]. However, in the present study, single heterozygous mice and MEFs ( Brca2 Ko/+ , Brca2 G25R/+ , and Palb2 Ko/+ ) did not reveal any overt defects.…”

Homologous DNA Repair:Safeguarding Genome Territories from Knives

“…a.) and functionally associates it with BRCA2 via its amino and carboxyl (6-90 a. a specifically Leu21 and 24) terminus respectively 13,15,17,[38][39][40] . Thus, the functional partnership of BRCA1 and BRCA2 in the DDR further elaborates the genetic similarity of either protein (Figure 2) 17,34 .…”