Gardberg Maria, Talvinen Kati, Kaipio Katja, Iljin Kristiina, Kampf Caroline, Uhlen Mathias, Carpén Olli
Department of Pathology, University of Turku and Turku University Central Hospital, Turku, Finland.
BMC Cell Biol. 2010 Jul 15;11:55. doi: 10.1186/1471-2121-11-55.
Diaphanous-related formins govern actin-based processes involved in many cellular functions, such as cell movement and invasion. Possible connections to developmental processes and cellular changes associated with malignant phenotype make them interesting study targets. In spite of this, very little is known of the tissue distribution and cellular location of any mammalian formin. Here we have carried out a comprehensive analysis of the formin family member formin -like 2 (FMNL2) in human tissues.
An FMNL2 antibody was raised and characterized. The affinity-purified FMNL2 antibody was validated by Western blotting, Northern blotting, a peptide competition assay and siRNA experiments. Bioinformatics-based mRNA profiling indicated that FMNL2 is widely expressed in human tissues. The highest mRNA levels were seen in central and peripheral nervous systems. Immunohistochemical analysis of 26 different human tissues showed that FMNL2 is widely expressed, in agreement with the mRNA profile. The widest expression was detected in the central nervous system, since both neurons and glial cells expressed FMNL2. Strong expression was also seen in many epithelia. However, the expression in different cell types was not ubiquitous. Many mesenchymal cell types showed weak immunoreactivity and cells lacking expression were seen in many tissues. The subcellular location of FMNL2 was cytoplasmic, and in some tissues a strong perinuclear dot was detected. In cultured cells FMNL2 showed mostly a cytoplasmic localization with perinuclear accumulation consistent with the Golgi apparatus. Furthermore, FMNL2 co-localized with F-actin to the tips of cellular protrusions in WM164 human melanoma cells. This finding is in line with FMNL2's proposed function in the formation of actin filaments in cellular protrusions, during amoeboid cellular migration.
FMNL2 is expressed in multiple human tissues, not only in the central nervous system. The expression is especially strong in gastrointestinal and mammary epithelia, lymphatic tissues, placenta, and in the reproductive tract. In cultured melanoma cells, FMNL2 co-localizes with F-actin dots at the tips of cellular protrusions.
Diaphanous相关的formin蛋白调控参与许多细胞功能（如细胞运动和侵袭）的基于肌动蛋白的过程。其与发育过程以及与恶性表型相关的细胞变化之间可能存在的联系，使其成为有趣的研究靶点。尽管如此，对于任何哺乳动物formin蛋白的组织分布和细胞定位却知之甚少。在此，我们对人组织中的formin家族成员formin样蛋白2（FMNL2）进行了全面分析。
制备并鉴定了一种FMNL2抗体。通过蛋白质免疫印迹法、Northern印迹法、肽竞争试验和小干扰RNA实验对亲和纯化的FMNL2抗体进行了验证。基于生物信息学的mRNA谱分析表明，FMNL2在人组织中广泛表达。在中枢和外周神经系统中观察到最高的mRNA水平。对26种不同人组织的免疫组织化学分析表明，FMNL2广泛表达，这与mRNA谱一致。在中枢神经系统中检测到最广泛的表达，因为神经元和神经胶质细胞均表达FMNL2。在许多上皮组织中也观察到强表达。然而，在不同细胞类型中的表达并非普遍存在。许多间充质细胞类型显示出弱免疫反应性，并且在许多组织中可见缺乏表达的细胞。FMNL2的亚细胞定位在细胞质中，并且在一些组织中检测到强的核周点状结构。在培养细胞中，FMNL2大多显示出细胞质定位，核周积累与高尔基体一致。此外，在WM164人黑色素瘤细胞中，FMNL2与F-肌动蛋白共定位于细胞突起的尖端。这一发现与FMNL2在变形虫样细胞迁移过程中在细胞突起中形成肌动蛋白丝的推测功能一致。
FMNL2在多种人组织中表达，不仅在中枢神经系统中。在胃肠道和乳腺上皮、淋巴组织、胎盘以及生殖道中表达尤其强烈。在培养的黑色素瘤细胞中，FMNL2与细胞突起尖端的F-肌动蛋白点共定位。
