The mechanism and function of ribosome-independent association of mRNA to the endoplasmic reticulum

人类基因组包含大约2.3万个基因，每个基因编码一种特定的蛋白质。这些基因的表达因细胞类型的不同而不同，并且可以受到包括环境提示在内的多种输入的调节。相当一部分基因(约占人类基因组的22%)编码蛋白质，这些蛋白质要么整合到细胞膜上，要么跨膜运输，从细胞中分泌出来。 基因表达始于细胞核，在那里包含基因的一段DNA首先被复制到信使RNA(MRNA)转录本中。然后，这些mRNAs从细胞核输出到细胞质，在那里蛋白质合成发生。虽然大多数mRNAs是由“游离”核糖体翻译成蛋白质的，但编码分泌或膜结合蛋白的转录本是由结合在内质网(ER)表面的核糖体翻译的，内质网是几乎所有细胞中都存在的一个大的网状细胞器。20世纪70年代的经典研究表明，这些mRNAs是通过在翻译过程中与核糖体结合而靶向并维持在内质网上的。然而，目前尚不清楚是否存在其他替代机制，以独立于核糖体或翻译来靶向和维持内质网上的这些mRNAs。使用新的方法，我们已经证明了许多编码分泌或膜结合蛋白的mRNAs可以靶向并维持在内质网上，即使在翻译被抑制或核糖体被分解时也是如此。此外，我们已经确定了可能与这种翻译非依赖性靶向有关的潜在的mRNA受体。在这份提案中，我将描述我们的计划，以确定哪些转录本使用这种替代的靶向/维持机制，确定RNA中存在的负责靶向的元件，以及确定内质网上与这些元件结合的假定的mRNA受体。通过这项工作，我希望对mRNA的胞质定位如何有助于细胞组织和分泌等各种基本细胞功能有更深入的了解。

The human genome contains about 23 thousand genes, each encoding a particular protein. The expression of these genes varies between cell type and can be modulated by a multitude of inputs including environmental cues. A substantial fraction of genes (~22% of the human genome) encode proteins that are either integrated into membranes or are transported across membranes to be secreted from the cell. Gene expression begins in the nucleus where a segment of DNA that contains the gene is first copied into messenger RNA (mRNA) transcripts. These mRNAs are then exported from the nucleus into the cytoplasm where protein synthesis takes place. Although most mRNAs are translated into proteins by "free" ribosomes, transcripts that encode secreted or membrane-bound proteins are translated by ribosomes bound to the surface of the endoplasmic reticulum (ER), a large reticular organelle found in almost all cells. Classic studies from the 1970s demonstrated that these mRNAs are targeted and maintained on the ER by associating to the ribosome during translation. It however was unclear whether additional alternative mechanisms existed to target and maintain these mRNAs on the ER independently of ribosomes or translation. Using novel approaches we have demonstrated that many mRNAs that encode secreted or membrane-bound proteins can target and be maintained on the ER even either when translation is inhibited or when ribosomes are disassembled. In addition, we have identified potential mRNA receptors that may be responsible for this translation-independent targeting. In this proposal I will describe our plans to identify which transcripts use this alternative targeting/maintenance mechanism, identify elements present within the RNA that are responsible for targeting, and identify the putative mRNA receptors on the ER that bind to these elements. Through this work, I hope to gain a deeper understanding of how the cytoplasmic localization of mRNA contributes to cellular organization and various basic cellular functions such as secretion.