Fracella Franco, Scholle Carl, Kallies Andreas, Häfker Thomas, Schröder Torsten, Rensing Ludger
Institute of Cell Biology, Biochemistry and Biotechnology, University of Bremen,PO Box 33 04 40, D-28334 Bremen,Germany.
Microbiology (Reading). 1997 Nov;143 ( Pt 11):3615-3624. doi: 10.1099/00221287-143-11-3615.
The constitutive and the heat-shock-induced expression of members of heat-shock protein families changed during vegetative development and conidiation of Neurospora crassa as determined by two-dimensional gel electrophoresis. Western blot and ELISA analyses revealed the highest amounts of the constitutive heat-shock protein 70 (HSC70) in conidiating aerial hyphae and dormant conidia. During conidial germination the amount of HSC70 decreased and subsequently increased during vegetative growth. Stationary mycelia and young aerial hyphae exhibited the lowest HSC70 level. The stationary-phase-dependent decrease in HSC70 was accompanied by a concomitant increase in its nuclear localization, whereas no significant changes in the amount of nuclear HSC70 were found during aerial hyphae development. The cAMP content during aerial hyphae development was inversely correlated with that of HSC70. To examine possible causal relations between HSC70 expression and cAMP content, the adenylate-cyclase-deficient mutant crisp (cr-1) was analysed, which exhibits low concentrations of endogenous cAMP. This mutant, however, showed a lower constitutive HSC70 level, compared to the bdA strain. Treatment of the bd strain and cr-1 mutant with 20 microM 8-bromo-cAMP did not result in significant changes of the constitutive HSC70 level, but in the level of heat-induced HSC/HSP70. In a developmental mutant (acon-2) which is defective in a differentiation step toward conidiation, the expression of HSC70 in aerial hyphae was delayed until the first proconidial chains were observed. It is concluded that the differential expression of HSC/HSP70 does not depend on different nuclear levels of HSC70 or on changes in cAMP concentrations, but rather on developmental genes controlling conidiation.
通过二维凝胶电泳确定，在粗糙脉孢菌的营养发育和分生孢子形成过程中，热休克蛋白家族成员的组成型表达和热休克诱导表达发生了变化。蛋白质免疫印迹和酶联免疫吸附测定分析表明，组成型热休克蛋白70（HSC70）在分生的气生菌丝和休眠分生孢子中的含量最高。在分生孢子萌发过程中，HSC70的含量下降，随后在营养生长过程中增加。静止菌丝体和幼嫩气生菌丝中HSC70水平最低。HSC70的静止期依赖性下降伴随着其核定位的相应增加，而在气生菌丝发育过程中，核HSC70的含量没有显著变化。气生菌丝发育过程中的cAMP含量与HSC70的含量呈负相关。为了研究HSC70表达与cAMP含量之间可能的因果关系，分析了腺苷酸环化酶缺陷型突变体crisp（cr-1），其内源cAMP浓度较低。然而，与bdA菌株相比，该突变体的组成型HSC70水平较低。用20 microM 8-溴-cAMP处理bd菌株和cr-1突变体，组成型HSC70水平没有显著变化，但热诱导的HSC/HSP70水平发生了变化。在一个在分生孢子形成的分化步骤中存在缺陷的发育突变体（acon-2）中，气生菌丝中HSC70的表达延迟，直到观察到第一批前分生孢子链。得出的结论是，HSC/HSP70的差异表达不依赖于HSC70的不同核水平或cAMP浓度的变化，而是依赖于控制分生孢子形成的发育基因。
