Copy number variation and gene expression

在许多动物和植物中，基因组的结构似乎非常浪费，大量的DNA显然没有功能，许多序列高度重复。除了基因组结构的复杂性之外，基因组的基因含量也存在差异。近年来，我们对这种基因拷贝数变异的理解，特别是对人类基因组的研究，取得了迅速的进展。对于大多数人类基因来说，我们都有两个拷贝——一个来自父母，一个来自父母。然而，对于某些基因，不同的人可能有不同的拷贝数，很明显，拷贝数很重要。恒河病毒阴性的人完全没有相应基因的拷贝，而恒河病毒阳性的人则有一个或两个拷贝。这些差异是母子不匹配的基础，可能导致“恒河猴宝宝”。同样，α -珠蛋白基因数量减少（通常每人4个拷贝）是全球常见血液疾病α -地中海贫血的基础。虽然自然选择通常确保变异在对其携带者产生负面影响时不会变得非常普遍，但很明显，在人类和许多其他物种中，有一些重要的基因表现出广泛的变异，种群中有许多不同的基因拷贝数。例如，编码唾液淀粉酶（负责消化食物中的淀粉）的基因以不同的数量存在——一些人只有两个拷贝，而另一些人则多达12个。有人认为，这种变异导致了唾液中淀粉酶产生的相应变异，从而导致了消化淀粉能力的变异。这是一个很吸引人的想法，但这是真的吗？更一般地说，基因数量的变化真的会导致基因功能的变化吗？如果是，又是如何导致的？实际上，情况很复杂。有两倍的基因并不一定意味着产生两倍的相关蛋白质——有一些例子表明，一个基因的额外拷贝可能会被使用，也可能不会被使用，或者只是部分活跃，这取决于它的精确位置，或者同一细胞中其他拷贝的数量。这个项目直接研究拷贝数的变化是否会导致基因表达的变化——如果是的话，以什么样的模式。为了做到这一点，我们将测量细胞和分泌物中的蛋白质水平，并询问它们与可变基因的拷贝数之间的关系。令人惊讶的是，计算基因组中DNA的拷贝数在技术上是困难的——事实上，对DNA进行测序比确切地知道有多少拷贝被测序要简单得多。显然，如果不能准确地测量拷贝数，那么对基因表达影响的推断也将是不准确的。因此，在这个项目中，准确地输入拷贝数变异是一个真正的挑战，以便（例如）一个测试可以清楚地区分一个个体是有6个还是7个可变基因拷贝。在进行这项工作时，我们小组的一个特别优势是在精确测量基因拷贝数方面的经验。该项目研究了人类基因组中三个数量可变基因的例子，它们在数量上表现出很大的差异；α -防御素（顾名思义，参与防御感染）通常在每人4到11个拷贝之间变化；唾液淀粉酶基因，每个人变异2-12个拷贝；最后，β -防御素DEFB109独立于α -防御素而变化（大多数人有2到7个拷贝），但也有一些拷贝因内部突变而失活——在这种情况下，基因表达可能不仅与拷贝总数有关，而且与能够制造活性蛋白质的拷贝数量有关。

In many animals and plants, the structure of the genome appears to be very wasteful, with large amounts of DNA apparently having no function, and many sequences being highly repeated. In addition to this complexity in genome structure, there is variation in the gene content of the genome. There have been rapid advances recently in our understanding of such variation in gene copy number, particularly in the study of the human genome. For most human genes, all of us have two copies - one from each parent. For some genes, however, different people can have different numbers of copies, and it is clear that the number of copies can matter. People who are Rhesus-negative have no copies at all of the corresponding gene, while Rhesus-positive individuals have one or two copies. These differences underlie the mother-child mismatch that can lead to a 'Rhesus baby' . Similarly, reduced numbers of the alpha-globin gene (usually four copies per person) are the basis of the globally common blood disease alpha-thalassaemia. Although natural selection generally ensures that variants do not get to be very common if they have negative effects on their carriers, it is clear that in humans and many other species, there are some important genes that show extensive variation, with many different gene copy numbers in the population. For example, the gene encoding the salivary amylase enzyme, responsible for digestion of starch in food, is present in variable numbers - some people have as few as two copies of this gene, while others can have as many as 12. It has been suggested that this variation leads to corresponding variation in the production of amylase in saliva, and so to variation in ability to digest starch. This is an attractive idea, but is it really true? More generally, does variation in gene number really lead to variation in gene function - and if so, how? In reality, the situation is complicated. Having twice as many genes does not necessarily mean that twice as much of the relevant protein is made - there are examples in which extra copies of a gene may or may not be used, or be only partially active, depending on its precise position, or on the number of other copies in the same cell. This project examines directly whether copy number variation leads to changes in gene expression - and if so in what patterns. To do this we will measure protein levels in cells and secretions, and ask how they relate to the copy number of variable genes. Surprisingly, counting copies of DNA in a genome is technically difficult - indeed, sequencing DNA is much more straightforward than knowing exactly how many copies have been sequenced. Clearly, if copy number is not being measured accurately, deductions about the effect on gene expression will also be inaccurate. It is therefore a real challenge in this project to type copy number variation accurately, so that (for example) a test can clearly distinguish whether an individual has 6 or 7 copies of a variable gene. One particular advantage our group has in undertaking this work is experience in the accurate measurement of gene copy number. The project examines three examples of variable-number genes from the human genome that show wide variation in number; the alpha-defensins (involved, as their name implies, in defence against infection) vary commonly between 4 and 11 copies per person; the salivary amylase gene, varying 2-12 copies per person; and finally, the beta-defensin DEFB109 varies independently of the alpha-defensins (between 2 and 7 copies in most people) but also has some copies that are inactivated by an internal mutation - in this case it is likely that gene expression will relate not just to the total number of copies, but the number of copies capable of making an active protein.