Tinsley J H, Lee I H, Minke P F, Plamann M
Department of Biology, Texas A&M University, College Station 77843-3258, USA.
Mol Gen Genet. 1998 Oct;259(6):601-9. doi: 10.1007/s004380050853.
The genes encoding actin and ARP3 in the filamentous fungus Neurospora crassa were cloned and sequenced. The actin structural gene is interrupted by four introns and encodes a polypeptide of 375 amino acids, which shows very high degree of identity with actin from other sources. N. crassa ARP3 is 439 amino acids in length and is 71% to 80% identical to ARP3s from five other organisms, while actin is 40% to 50% identical to these same ARP3s. Transcript levels for actin and ARP3 decrease upon induction of asexual development (i.e. conidiation) and subsequently increase slightly when conidia are being formed. A concentration of filamentous actin is typically seen at sites of growth in eukaryotic organisms and, using indirect immunofluorescence, we showed that filamentous actin is localized primarily to hyphal tips in N. crassa. To determine if the level of actin increases in response to an increase in the number of growth sites and in the area of the growing surface, we used the temperature-sensitive mutants cot-1 and mcb. Growth of the cot-1 and mcb mutants at restrictive temperature induces hyphal tip formation and a loss of growth polarity, respectively. Unexpectedly, almost no increase in actin levels is observed following a > 20-fold increase in the number of hyphal tips or an increase in the area of the growing surface resulting from a loss of growth polarity. The results suggest that the level of actin monomers within N. crassa hyphae is sufficient to accommodate the need for additional actin patches and filaments that arises when the number of hyphal tips and the area of growing surface per unit length of hypha greatly exceeds that in wild-type.
对丝状真菌粗糙脉孢菌中编码肌动蛋白和ARP3的基因进行了克隆和测序。肌动蛋白结构基因被四个内含子打断，编码一个由375个氨基酸组成的多肽，该多肽与其他来源的肌动蛋白具有非常高的同源性。粗糙脉孢菌的ARP3长度为439个氨基酸，与其他五种生物的ARP3同源性为71%至80%，而肌动蛋白与这些相同的ARP3同源性为40%至50%。在无性发育（即分生孢子形成）诱导后，肌动蛋白和ARP3的转录水平下降，随后在分生孢子形成时略有增加。在真核生物的生长部位通常可见丝状肌动蛋白的聚集，并且通过间接免疫荧光，我们表明丝状肌动蛋白主要定位于粗糙脉孢菌的菌丝尖端。为了确定肌动蛋白水平是否随着生长位点数量的增加和生长表面面积的增加而增加，我们使用了温度敏感突变体cot-1和mcb。cot-1和mcb突变体在限制温度下生长分别诱导菌丝尖端形成和生长极性丧失。出乎意料的是，在菌丝尖端数量增加>20倍或由于生长极性丧失导致生长表面面积增加后，几乎没有观察到肌动蛋白水平的增加。结果表明，粗糙脉孢菌菌丝内肌动蛋白单体的水平足以满足当菌丝尖端数量和每单位长度菌丝的生长表面面积大大超过野生型时对额外肌动蛋白斑块和细丝的需求。
