Differential Expression of BARD1 Isoforms in Melanoma

Abstract

Melanoma comprises <5% of cutaneous malignancies, yet it causes a significant proportion of skin cancer-related deaths worldwide. While new therapies for melanoma have been developed, not all patients respond well. Thus, further research is required to better predict patient outcomes. Using long-range nanopore sequencing, RT-qPCR, and RNA sequencing analyses, we examined the transcription of BARD1 splice isoforms in melanoma cell lines and patient tissue samples. Seventy-six BARD1 mRNA variants were identified in total, with several previously characterised isoforms (γ, φ, δ, ε, and η) contributing to a large proportion of the expressed transcripts. In addition, we identified four novel splice events, namely, Δ(E3_E9), ▼(i8), IVS10+131▼46, and IVS10▼176, occurring in various combinations in multiple transcripts. We found that short-read RNA-Seq analyses were limited in their ability to predict isoforms containing multiple non-contiguous splicing events, as compared to long-range nanopore sequencing. These studies suggest that further investigations into the functional significance of the identified BARD1 splice variants in melanoma are warranted.

Keywords: BARD1; RNA-Seq; melanoma; nanopore sequencing.

Figure 1

Schematic diagram of BARD1 and identified isoforms (selected). (A) Structure of full-length BARD1 (BARD1-FL) and six abundantly expressed alternative isoforms (γ, φ, δ, ε, η, and Δ(E3_E9)). (B) Structure of identified splicing events. (C) Presence of identified isoforms as detected using various sequencing techniques. (nd)- not done.

Figure 2

Proportion of BARD1 transcript isoforms quantified using nanopore sequencing. The first three bars correspond to the New Zealand melanoma (NZM) cell lines, and the last bar corresponds to the non-malignant (melanocyte) control.