Soreq Lilach, Gilboa-Geffen Adi, Berrih-Aknin Sonia, Lacoste Paul, Darvasi Ariel, Soreq Eyal, Bergman Hagai, Soreq Hermona
Department of Physiology, The Hebrew University, Hadassah Medical School, Jerusalem, Israel.
PLoS One. 2008 Jun 11;3(6):e2392. doi: 10.1371/journal.pone.0002392.
The vast majority of human genes (>70%) are alternatively spliced. Although alternative pre-mRNA processing is modified in multiple tumors, alternative hyper-splicing signatures specific to particular tumor types are still lacking. Here, we report the use of Affymetrix Human Exon Arrays to spot hyper-splicing events characteristic of myasthenia gravis (MG)-thymoma, thymic tumors which develop in patients with MG and discriminate them from colon cancer changes.
METHODOLOGY/PRINCIPAL FINDINGS: We combined GO term to parent threshold-based and threshold-independent ad-hoc functional statistics with in-depth analysis of key modified transcripts to highlight various exon-specific changes. These denote alternative splicing in MG-thymoma tumors compared to healthy human thymus and to in-house and Affymetrix datasets from colon cancer and healthy tissues. By using both global and specific, term-to-parent Gene Ontology (GO) statistical comparisons, our functional integrative ad-hoc method allowed the detection of disease-relevant splicing events.
CONCLUSIONS/SIGNIFICANCE: Hyper-spliced transcripts spanned several categories, including the tumorogenic ERBB4 tyrosine kinase receptor and the connective tissue growth factor CTGF, as well as the immune function-related histocompatibility gene HLA-DRB1 and interleukin (IL)19, two muscle-specific collagens and one myosin heavy chain gene; intriguingly, a putative new exon was discovered in the MG-involved acetylcholinesterase ACHE gene. Corresponding changes in spliceosome composition were indicated by co-decreases in the splicing factors ASF/SF(2) and SC35. Parallel tumor-associated changes occurred in colon cancer as well, but the majority of the apparent hyper-splicing events were particular to MG-thymoma and could be validated by Fluorescent In-Situ Hybridization (FISH), Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and mass spectrometry (MS) followed by peptide sequencing. Our findings demonstrate a particular alternative hyper-splicing signature for transcripts over-expressed in MG-thymoma, supporting the hypothesis that alternative hyper-splicing contributes to shaping the biological functions of these and other specialized tumors and opening new venues for the development of diagnosis and treatment approaches.
绝大多数人类基因（>70%）存在可变剪接。尽管可变前体mRNA加工在多种肿瘤中会发生改变，但仍缺乏特定肿瘤类型特有的可变超剪接特征。在此，我们报告使用Affymetrix人类外显子芯片来发现重症肌无力（MG）-胸腺瘤（一种在MG患者中发生的胸腺肿瘤）的超剪接事件特征，并将其与结肠癌的变化区分开来。
方法/主要发现：我们将基于GO术语到父阈值和阈值无关的特设功能统计与对关键修饰转录本的深入分析相结合，以突出各种外显子特异性变化。这些变化表明与健康人胸腺以及来自结肠癌和健康组织的内部和Affymetrix数据集相比，MG-胸腺瘤肿瘤中存在可变剪接。通过使用全局和特定的、术语到父基因本体（GO）的统计比较，我们的功能整合特设方法能够检测与疾病相关的剪接事件。
结论/意义：超剪接转录本涵盖多个类别，包括致癌性ERBB4酪氨酸激酶受体和结缔组织生长因子CTGF，以及免疫功能相关的组织相容性基因HLA-DRB1和白细胞介素（IL）19、两种肌肉特异性胶原蛋白和一种肌球蛋白重链基因；有趣的是，在MG相关的乙酰胆碱酯酶ACHE基因中发现了一个推定的新外显子。剪接因子ASF/SF(2)和SC35的共同减少表明剪接体组成发生了相应变化。结肠癌中也发生了与肿瘤相关的平行变化，但大多数明显的超剪接事件是MG-胸腺瘤特有的，并且可以通过荧光原位杂交（FISH）、逆转录-聚合酶链反应（RT-PCR）和质谱（MS）以及随后的肽测序进行验证。我们的发现证明了MG-胸腺瘤中过表达的转录本具有特定的可变超剪接特征，支持了可变超剪接有助于塑造这些及其他特殊肿瘤的生物学功能这一假设，并为诊断和治疗方法的开发开辟了新途径。
