因為遺傳編碼的多義形成61種密碼子編碼20種胺基酸。編碼相同胺基酸的密碼子稱為同義密碼子。在所有報導過的基因組中，同義密碼子使用頻率並不相等的，這種現象稱為密碼子使用偏移。密碼子的使用偏移如何去調控人類的基因表達目前尚未完全了解。鑒於超過200種細胞類型，每種細胞類型都有不同的生理功能和型態。因此我假設至少有一部份透過細胞特異性的密碼子使用偏移來做基因調控。為了證明假設，我設計幾個具有不同密碼子使用模式的螢火蟲熒光素酶(FLUC)報告基因去測量在三種不同人類細胞中的冷光活性比。在人類胰腺癌細胞與人胚胎腎細胞中觀察到不同的冷光活性比。接著通過有限的胰蛋白酶消化實驗進一步檢查結果。然而這三種不同密碼使用模式在任一種細胞系中對胰蛋白酶的敏感性並沒有顯著差異。出乎意料的是，兩種靶向螢火蟲熒光素酶蛋白不同區域的單株抗體僅在人類胰腺癌細胞中對野生型螢火蟲熒光素酶(FLUC)表現出不同的敏感性。此結果增加當特定胺基酸由非最佳密碼子組成時具有高頻率解碼錯誤且跟細胞特異性相關的可能性。因此更進一步將實驗側重於密碼子解碼錯誤發生的位置識別。通過在螢火蟲熒光素酶(FLUC)的C端添加FLAG標籤去量化沒有密碼子影響下的螢火蟲熒光素酶(FLUC)蛋白表達量水平,我驚訝地觀察到與FLAG抗體在人類胰腺癌細胞(PANC1 cell)偵測到蛋白表現量水平相比，兩種靶向螢火蟲熒光素酶(FLUC)編碼區的單株抗體所偵測到的蛋白表現量是低估。總結而論，我的數據支持假設為密碼子使用偏移具有細胞特異性以適應蛋白質表達的需要。另外，我的數據也表明密碼子的使用偏移可能會在特定人類細胞中引起轉譯錯誤解碼。

Due to the degeneracy of genetic codes, sixty-one codons encode twenty amino acids. The codons that code for the same amino acid are called synonymous codons. In all reported genomes, the usage frequency of synonymous codons is not equal, a phenomenon termed codon usage bias. How does codon usage bias regulate gene expression in human cells is not completely understood. Given there are more than two hundred cell types that each has distinct physiological function and morphology, I hypothesize that cell type-specific gene regulation may be, at least in part, mediated by the cell type-specific codon usage bias. To test this, I designed firefly luciferase (FLUC) reporters with different codon usage patterns to measure their specific activity in three different human cell lines. I observed that the specific activity of the codon variant FLUCs in PANC1 and K562 cells are different while the specific activity of the FLUCs show no significant difference in HEK293T cells. These results are further examined by limited trypsin digestion. However, there is no significant difference of trypsin sensitivity of these three codon variants in either cell types. Unexpectedly, two monoclonal FLUC antibodies
that target different regions of FLUC show varied sensitivity to wildtype FLUC only in PANC1 cells. This result raises the possibility that certain residues with non-optimal codons may be decoded erroneously in high frequency and in a cell type-specific manner. By introducing a FLAG tag in the C-termimal of the codon variant FLUCs to quantify FLUC protein levels without codon effect, I observed that, surprisingly, both monoclonal antibodies that target FLUC coding region underestimate protein levels compared to the FLAG antibody detected ones in PANC1 cells. Together, my data support the hypothesis that there existing a cell-type specific codon bias to adapt to protein expression need. In addition, my data suggests that codon bias might introduce translational misdecoding in specific human cells.

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