Lu Hong, Li Liyuan, Watson Edmond R, Williams Robert W, Geisert Eldon E, Jablonski Monica M, Lu Lu
Department of Ophthalmology, Hamilton Eye Institute, University of Tennessee Health Science Center, Memphis, TN, USA.
Mol Vis. 2011;17:2455-68. Epub 2011 Sep 22.
To use a systems genetics approach to construct and analyze co-expression networks that are causally linked to mutations in a key pigementation gene, tyrosinase-related protein 1 (Tyrp1), that is associated both with oculocutaneous albinism type 3 (OCA3) in humans and with glaucoma in mice.
Gene expression patterns were measured in whole eyes of a large family of BXD recombinant inbred (RI) mice derived from parental lines that encode for wildtype (C57BL/6J) and mutant (DBA/2J) Tyrp1. Protein levels of Tyrp1 were measured in whole eyes and isolated irides. Bioinformatics analyses were performed on the expression data along with our archived sequence data. Separate data sets were generated which were comprised of strains that harbor either wildtype or mutant Tyrp1 and each was mined individually to identify gene networks that covary significantly with each isoform of Tyrp1. Ontology trees and network graphs were generated to probe essential function and statistical significance of covariation. Genes with strong covariance in wildtype mice were assembled into genome-wide heatmaps for cohorts carrying either wildtype or mutant Tyrp1.
Single nucleotide polymorphism (SNP) analysis verified the presence of the Tyrp1b mutation in the Tyrp1 gene. Message levels were greater in BXD strains with the mutant Tyrp1. Interval mapping of these BXD mice revealed a strong expression quantitative trait locus (eQTL) on Chr 4 at the location of the gene itself. Composite mapping revealed a suggestive eQTL on Chr 9 at the location of myosin-Va (Myo5a), mutations in which are known as dilute. Enriched biologic processes associated with wildtype Tyrp1 included pigmentation, melanin biosynthetic process, and mesenchymal cell development, while associations with the mutant gene included categories of neural crest cell development, protein metabolic processes and glycoprotein metabolic processes. Genome-wide heatmaps revealed strong candidate cis-eQTLs on Chr 4 at Tyrp1 and on Chr 9 at Myo5a in all mice. In the wildtype data set, Tyrp1 was an upstream regulator of six pigmentation and two mesenchyme genes. In addition, five genes, including Tyrp1, were at least partially regulated by Myo5a. Analyses of the strains harboring the mutant gene revealed significant loss of correlation to traditional genes and gain of correlation to genes with little or no functional relationship.
These findings indicate that the Tyrp1(b) mutation modifies the pathways and gene networks in which Tyrp1 functions. Our results also indicate direct and indirect regulatory control of Tyrp1 and other pigmentation and mesenchymal genes by Myo5a. Lastly, we find that the mutations reduce the ability of Tyrp1 to regulate expression of other genes that participate in pigmentation metabolism.
采用系统遗传学方法构建并分析与关键色素沉着基因酪氨酸酶相关蛋白1（Tyrp1）突变有因果关联的共表达网络。Tyrp1与人类3型眼皮肤白化病（OCA3）以及小鼠青光眼均相关。
在一个源自编码野生型（C57BL/6J）和突变型（DBA/2J）Tyrp1的亲本品系的大型BXD重组近交（RI）小鼠家族的全眼中测量基因表达模式。在全眼和分离出的虹膜中测量Tyrp1的蛋白水平。对表达数据以及我们存档的序列数据进行生物信息学分析。生成了单独的数据集，其中一组包含携带野生型Tyrp1的品系，另一组包含携带突变型Tyrp1的品系，分别对每组进行挖掘以识别与Tyrp1各同种型显著共变的基因网络。生成本体树和网络图以探究共变的基本功能和统计显著性。将野生型小鼠中具有强共变性的基因整合到携带野生型或突变型Tyrp1的群体的全基因组热图中。
单核苷酸多态性（SNP）分析证实了Tyrp1基因中存在Tyrp1b突变。携带突变型Tyrp1的BXD品系中的信使水平更高。对这些BXD小鼠进行区间定位发现在4号染色体上该基因本身的位置有一个强表达数量性状基因座（eQTL）。复合定位发现在9号染色体上肌球蛋白-Va（Myo5a）所在位置有一个暗示性eQTL，其突变被称为稀释。与野生型Tyrp1相关的富集生物学过程包括色素沉着、黑色素生物合成过程和间充质细胞发育，而与突变基因相关的类别包括神经嵴细胞发育、蛋白质代谢过程和糖蛋白代谢过程。全基因组热图显示在所有小鼠中，4号染色体上Tyrp1位置以及9号染色体上Myo5a位置存在强候选顺式eQTL。在野生型数据集中，Tyrp1是六个色素沉着基因和两个间充质基因的上游调节因子。此外，包括Tyrp1在内的五个基因至少部分受Myo5a调节。对携带突变基因的品系进行分析发现与传统基因的相关性显著丧失，与功能关系很少或没有功能关系的基因的相关性增加。
这些发现表明Tyrp1(b)突变改变了Tyrp1发挥功能的途径和基因网络。我们的结果还表明Myo5a对Tyrp1以及其他色素沉着和间充质基因有直接和间接的调控作用。最后，我们发现这些突变降低了Tyrp1调节参与色素沉着代谢的其他基因表达的能力。
