The F-box protein, FBXO7, is required to maintain chromosome stability in humans.

Despite the high morbidity and mortality rates associated with colorectal cancer (CRC), the aberrant genes and mechanisms driving CRC pathogenesis remain poorly understood. Chromosome instability (CIN), or ongoing changes in chromosome numbers, is a predominant form of genome instability associated with ~85% of CRCs, suggesting it may be a key mechanism driving CRC oncogenesis. CIN enables the acquisition of copy number alterations conferring selective growth, proliferation and survival advantages that promote cellular transformation. Despite these associations, the aberrant genes underlying CIN remain largely unknown. Candidate CIN gene FBXO7 encodes an F-box protein, a subunit of the SKP1-CUL1-FBOX (SCF) complex that confers substrate specificity to the complex and targets proteins for subsequent degradation by the 26S proteasome. Recently, the genes encoding the three core SCF complex members were identified as CIN genes; however, it is unknown whether F-box proteins exhibit similar integral roles in maintaining chromosome stability. Using short- small interfering RNA (siRNA) and long- (CRISPR/Cas9) term approaches, we show that reduced FBXO7 expression induces CIN in various colonic epithelial cell contexts, whereas FBXO7 knockout clones also exhibit hallmarks associated with cellular transformation, namely increased clonogenic and anchorage-independent growth. Collectively, these data demonstrate that FBXO7 is required to maintain genome stability identifying FBXO7 a novel CIN gene whose reduced expression may contribute to CRC development and progression.

A Comprehensive Assessment of Genetic and Epigenetic Alterations Identifies Frequent Variations Impacting Six Prototypic SCF Complex Members.

The SKP1, CUL1, F-box protein (SCF) complex represents a family of 69 E3 ubiquitin ligases that poly-ubiquitinate protein substrates marking them for proteolytic degradation via the 26S proteasome. Established SCF complex targets include transcription factors, oncoproteins and tumor suppressors that modulate cell cycle activity and mitotic fidelity. Accordingly, genetic and epigenetic alterations involving SCF complex member genes are expected to adversely impact target regulation and contribute to disease etiology. To gain novel insight into cancer pathogenesis, we determined the prevalence of genetic and epigenetic alterations in six prototypic SCF complex member genes (SKP1, CUL1, RBX1, SKP2, FBXW7 and FBXO5) from patient datasets extracted from The Cancer Genome Atlas (TCGA). Collectively, ~45% of observed SCF complex member mutations are predicted to impact complex structure and/or function in 10 solid tumor types. In addition, the distribution of encoded alterations suggest SCF complex members may exhibit either tumor suppressor or oncogenic mutational profiles in a cancer type dependent manner. Further bioinformatic analyses reveal the potential functional implications of encoded alterations arising from missense mutations by examining predicted deleterious mutations with available crystal structures. The SCF complex also exhibits frequent copy number alterations in a variety of cancer types that generally correspond with mRNA expression levels. Finally, we note that SCF complex member genes are differentially methylated across cancer types, which may effectively phenocopy gene copy number alterations. Collectively, these data show that SCF complex member genes are frequently altered at the genetic and epigenetic levels in many cancer types, which will adversely impact the normal targeting and timely destruction of protein substrates, which may contribute to the development and progression of an extensive array of cancer types.

Reduced SKP2 Expression Adversely Impacts Genome Stability and Promotes Cellular Transformation in Colonic Epithelial Cells.

Despite the high morbidity and mortality rates associated with colorectal cancer (CRC), the underlying molecular mechanisms driving CRC development remain largely uncharacterized. Chromosome instability (CIN), or ongoing changes in chromosome complements, occurs in ~85% of CRCs and is a proposed driver of cancer development, as the genomic changes imparted by CIN enable the acquisition of karyotypes that are favorable for cellular transformation and the classic hallmarks of cancer. Despite these associations, the aberrant genes and proteins driving CIN remain elusive. SKP2 encodes an F-box protein, a variable subunit of the SKP1-CUL1-F-box (SCF) complex that selectively targets proteins for polyubiquitylation and degradation. Recent data have identified the core SCF complex components (SKP1, CUL1, and RBX1) as CIN genes; however, the impact reduced SKP2 expression has on CIN, cellular transformation, and oncogenesis remains unknown. Using both short- small interfering RNA (siRNA) and long-term (CRISPR/Cas9) approaches, we demonstrate that diminished SKP2 expression induces CIN in both malignant and non-malignant colonic epithelial cell contexts. Moreover, temporal assays reveal that reduced SKP2 expression promotes cellular transformation, as demonstrated by enhanced anchorage-independent growth. Collectively, these data identify SKP2 as a novel CIN gene in clinically relevant models and highlight its potential pathogenic role in CRC development.

Reduced RBX1 expression induces chromosome instability and promotes cellular transformation in high-grade serous ovarian cancer precursor cells.

Despite high-grade serous ovarian cancer (HGSOC) being the most common and lethal gynecological cancer in women, the early etiological events driving disease development remain largely unknown. Emerging evidence now suggests that chromosome instability (CIN; ongoing changes in chromosome numbers) may play a central role in the development and progression of HGSOC. Importantly, genomic amplification of the Cyclin E1 gene (CCNE1) contributes to HGSOC pathogenesis in ~20% of patients, while Cyclin E1 overexpression induces CIN in model systems. Cyclin E1 levels are normally regulated by the SCF (SKP1-CUL1-FBOX) complex, an E3 ubiquitin ligase that includes RBX1 as a core component. Interestingly, RBX1 is heterozygously lost in ~80% of HGSOC cases and reduced expression corresponds with worse outcomes, suggesting it may be a pathogenic event. Using both short (siRNA) and long (CRISPR/Cas9) term approaches, we show that reduced RBX1 expression corresponds with significant increases in CIN phenotypes in fallopian tube secretory epithelial cells, a cellular precursor of HGSOC. Moreover, reduced RBX1 expression corresponds with increased Cyclin E1 levels and anchorage-independent growth. Collectively, these data identify RBX1 as a novel CIN gene with pathogenic implications for HGSOC.