腦指蛋白 (Brain Finger Protein, BFP)，又名為Znf179，屬於環指蛋白 (RING finger) 家族成員之一。Znf179基因會伴隨著胚胎腦部發育過程而表現，顯示在腦部發育及維持腦部功能其可能扮演重要的角色。在實驗室先前的研究發現，P19細胞分化成神經細胞的過程中，Znf179基因表現會隨著神經細胞的分化逐漸增加，而在P19細胞進行分化的過程中抑制Znf179的表現會明顯影響到P19細胞的神經分化，顯示Znf179基因的確在神經細胞分化的過程扮演重要的角色，但到目前為止對於Znf179在細胞中的分子作用機制仍不清楚，因此我的研究主要是藉由酵母菌雙雜合系統 (Yeast two-hybrid)，試著從老鼠腦部基因庫中找出可能與Znf179基因有結合作用的蛋白質，希望藉此推測出Znf179基因在細胞內可能的分子作用機制。我們在酵母菌雙雜合系統實驗中發現，Znf179基因會與早幼粒細胞白血病鋅指蛋白 (Promyelocytic leukemia Zinc-Finger, PLZF) 結合，我們也進一步利用免疫沉澱法 (Immunoprecipitation) 證明兩個蛋白間的確有交互作用，並且PLZF蛋白質可能是透過C端鋅指功能區的前兩個鋅指與Znf179產生結合。從免疫螢光染色 (Immunofluorescence) 結果我們也發現，當PLZF與Znf179同時存在的情況下，PLZF會改變Znf179在細胞中分佈的位置，以上的實驗結果都顯示在細胞中這兩個蛋白質確實會互相結合。已知PLZF是個轉錄抑制因子，會吸引調控轉錄作用的協同抑制因子 (transcriptional co-repressor)，例如：N-COR、SMART、Histone deacetylase (HDAC)，形成複合體，達到抑制基因的轉錄作用。為了探討Znf179基因是否影響PLZF的轉錄調控能力，我們共同轉殖Flag-PLZF及EGFP-Znf179或控制組質體，結果我們意外地發現，在Znf179存在的情況下PLZF的蛋白質表現量有明顯增加的現象，但在RNA的層面則沒有明顯的影響，因此在未來的研究上會繼續探討Znf179對於PLZF在蛋白質的轉譯及穩定性上的調控機制。

Znf179 is a member of the RING finger protein family. During embryogenesis, Znf179 is restrictedly expressed in brain region, suggesting a potential role in neuronal development. Previous studies in our laboratory have demonstrated that the expression of Znf179 is remarkably increased in the retinoic acid-induced neuronal differentiated P19 cells and Znf179 knock-down significantly attenuates neuronal differentiation. To investigate the molecular mechanisms of Znf179 function, we performed yeast two-hybrid screen to identify the Znf179 interacting proteins. In the experiments, promyelocytic leukemia zinc-finger (PLZF) was identified as a novel Znf179 interacting protein. The results of immunoprecipitation assay also confirmed the interaction between PLZF and Znf179 in vivio. Furthermore, we have demonstrated that PLZF interacts with Znf179 via its two zinc-finger domains, which is located between amino acids 398 and 455. Using immunofluorescence analysis, we observed that the cellular localization of Znf179 was changed by co-expressing PLZF. Taken together, these results indicate that the two proteins dose indeed interact with each other in vivo. PLZF has been found to regulate transcription through recruitment of nuclear receptor corepressors (N-CoR or SMRT) / histone deacetylase (HDAC) complexes via its POZ domain. To investigate whethere Znf179 affects transcriptional regulation ability of PLZF, COS-1 cells were co-transfected with HA-PLZF and EGFP-Znf179 or control plasmid. Unexpectedly, we found that exogenously expressing Znf179 caused a 5-fold increase in HA-PLZF protein abundance but not at Flag-PLZF RNA level. Further studies will be performed to examine the molecular mechanisms of Znf179 functions on PLZF protein regulation.

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