Is Damage and Repair of DNA Required for Differentiation?

一项发现HL60细胞在紫外线(UV)照射下发生终末分化的发现引发了一个问题：当紫外线形成的T-T二聚体被修复时，核DNA中单链刹车(SSB)的瞬时形成是否诱导了细胞分化？因此，我们检验了“DNA中SSB的形成及其修复是细胞分化和衰老所必需的”这一假设的有效性。利用外周血淋巴细胞进行了一系列实验。为了进行实验，开发了一种原位缺口平移方法，以组织化学方法在每个细胞核中定位SSB。外周血淋巴细胞暴露于紫外线照射后约2小时形成SSB，此后逐渐下降。T-T二聚体在照射后即刻出现频率最高，并逐渐降低。这些发现，再加上先前的发现，当淋巴细胞暴露在紫外线下时，淋巴细胞产生了一系列在有丝分裂原刺激细胞时出现的蛋白质，提示SSB的形成和修复与细胞的分化有关。为了检验这种现象是否也发生在体内，研究人员将小鼠的耳朵暴露在紫外线下，并进行了类似于对淋巴细胞进行的研究。研究发现，表皮中的基底层细胞具有修复SSB的能力，而棘细胞层中的细胞则失去了修复SSB的能力。在基底细胞中，有已存在的SSB和新形成的SSB。前者是DNA聚合酶不能到达的，只有后者得到了修复。这些发现表明，在一个给定的核中，存在两个结构域，一个是SSB被修复并从DNA转录到hnRNA的区域，另一个是SSB保留而不发生转录的区域。

A finding that HL60 cells when exposed to ultraviolet light (UV) underwent terminal differentiation has led to a question that does transient formation of single stranded brakes (SSB) in nuclear DNA as T-T dimers formed by UV are repaired induced the cell differentiation? Hence, we examined the validity of hypothesis that "Formation of SSB in DNA and their repair is required for cell differentiation and aging". A series of experiments were carried out using peripheral lymphocytes. To carry out the experiments, an in situ nick translation method was developed in order to localize SSB histochemically in each cell nuclei. When peripheral lymphocytes were exposed to UV, SSB was formed about 2 hours after the UV irradiation and the SSB gradually decreased thereafter. The T-T dimers were most frequent immediately after the irradiation and gradually decreased. These findings together with the previous finding that when the lymphocytes were exposed to UV, the lymphocytes produced a series of proteins which appear when the cells were stimulated with mitogen suggested that the SSB formation and their repair are associated with the cell differentiation. To examine whether the phenomenon also takes place in in vivo, ears of mice were exposed to UV and studies similar to that done on the lymphocytes were carried out. It was found that the basal cells in epidermis has an ability to repair SSB whereas cells in prickle cell layers has lost the ability. In the basal cells, there were pre-existed SSB and newly formed SSB. The former are inaccessible to DNA polymerase and only the latter were repaired. These finding suggested that in a given nuclei there are two domains, one where SSB are repaired and transcription from DNA to hnRNA takes place and another where SSB remains and no transcription takes place.