Purification and Partial Amino Acid Sequence of the Major 70,000-Dalton Heat Shock Protein in Neurospora crassa

Fracella, Franco; Mohsenzadeh, Saadat; Rensing, Ludger

doi:10.1006/emyc.1993.1035

Cited by 15 publications

(24 citation statements)

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“…The amino acid sequence includes one Trp, four Cys, and five His residues; 28% hydrophobic residues; and 15 and 12%, respectively, acidic and basic residues. Recently the purification and the amino acid sequence of the first 53 residues of the major 70-kDa HSP of N. crassa were reported (10). The published amino acid sequence is in complete agreement with that deduced by us for this part of the polypeptide.…”

Section: And Petunia (47)-as Well As In the Caenorhabditis Elegans Hssupporting

confidence: 81%

The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member

1995

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The gene encoding the major heat shock-inducible member of the HSP70 family of Neurospora crassa was cloned and characterized. The 5 nontranscribed region shows the presence of consensus sequence motifs resembling the classical heat shock elements found in many heat shock-responsive eukaryotic promoters, as well as metal-responsive-element sequences. The coding region of the gene contains four introns with boundaries and internal consensus motifs typical of genes of filamentous fungi. None of the other stress-inducible hsp70 genes of fungal origin have, so far, been reported to contain introns. The sequence adjoining the transcriptional initiation zone shows the presence of prominent CT-rich stretches, characteristic of highly expressed fungal genes. The deduced amino acid sequence corresponds to a 646-residue polypeptide, with a calculated molecular mass of 70,561 Da and an average pI of 6.01, exhibiting strong sequence homology with many other eukaryotic HSP70s, with typical HSP70 family signatures 1 and 2 and a bipartite nuclear targeting sequence. Experiments with primer extension revealed the presence of one minor and two major transcriptional start sites. This gene, designated hsps-1, was mapped to a locus on the left arm of linkage group II, in close proximity to the AR-30 translocation breakpoint.The heat shock response is a fundamental attribute of living organisms, enabling them to withstand sudden environmental changes. The rapid synthesis of a specific complement of heat shock proteins (HSPs), or stress-responsive proteins, following hyperthermal injury or application of physiological or metabolic stress is recognized as an effective, protective strategy for ensuring cellular survival (32). In eukaryotic organisms, HSPs with a large molecular size (70 to 110 kDa) appear to be implicated in several functions critical for the maintenance of the integrity of structural proteins and cellular enzymes under unfavorable conditions.The stress-inducible HSP70 members compose one of the most highly conserved protein and gene families encountered throughout the biological world (4). The importance of the role of the stress-70 class of proteins as molecular chaperones of protein folding and oligomeric assembly, during heat shock as well as in normal cellular physiology, is beginning to be appreciated, and there is beginning to be a greater understanding of their structure-function relationships (13). The hsp70 multigene family of yeast, comprising eight members-including constitutive as well as stress-inducible ones-has been investigated intensively (6). In contrast, very little information is available on the heat shock genes and proteins of the filamentous fungi. Information derived from recent phylogenetic studies at the molecular level provides strong support for a close evolutionary linkage between the fungal and animal kingdoms (2). Therefore, elucidation of the evolutionary basis of defensive strategies elaborated by the filamentous fungi should provide invaluable genetic tools for exploration of similar ...

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Section: And Petunia (47)-as Well As In the Caenorhabditis Elegans Hssupporting

confidence: 81%

The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member

1995

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“…The blots were blocked with TPBS for 1 h at 25°C. After removal of the blocking solution, primary antibody (affinity-purified polyclonal rabbit anti-HSP70 against the major Neurospora HSP70; Fracella et al, 1993) diluted in TPBS (1:1000), was added and left for 1 h at 25°C. Subsequently, the blots were washed with TPBS.…”

Section: Western Blot Analysismentioning

confidence: 99%

“…Rabbits were immunized with primary injections of 30 µg of dialized and lyophilized isolated HSP70 from N. crassa (Fracella et al, 1993) in 0.25 ml PBS in a 1:1 mixture with Freund's complete adjuvant (Calbiochem, Bad Soden) distributed to two subcutaneous sites on the back. Booster injections of 15 µg of HSP70 were given at 14-day intervals after primary injection in an identical manner except that incomplete Freund's adjuvant (Calbiochem) was used.…”

Section: Antibody Preparationmentioning

confidence: 99%

Temperature Adaptation of House Keeping and Heat Shock Gene Expression inNeurospora crassa

Mohsenzadeh

Saupe-Thies

Steier

et al. 1998

Fungal Genetics and Biology

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“…These hyphae differentiate into aerial hyphae capable of developing macroconidia (Springer 8i Yanofsky, 1989) directed by a circadian clock and in response to environmental stress, such as nutrient deprivation, desiccation, oxygen (Nelson et al , 1975), heat stress (Rensing, 1993) or light exposure (Lauter & RUSSO, 1991). When dormant conidia are activated by hydration in the presence of suitable nutrients, they germinate, again develop a mycelium and thus complete a new vegetative cycle.…”

Section: Discussionmentioning

confidence: 99%

Differential expression of glucose-regulated (grp78) and heat-shock-inducible (hsp70) genes during asexual development of Neurospora crassa

et al. 1998

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The expression of a glucose-regulated gene (grp78) changes significantly during the vegetative life cycle of Neurospora crassa: the amounts of grp78 mRNA are low in dormant conidia, increase during germination and exponential growth, decline in young aerial hyphae and reach a maximum in late (15-18 h) aerial hyphae. Heat shock (30 min at 45 "C) elevated the mRNA level of this gene especially in early aerial hyphae, whereas no increase above the high constitutive amount was found after heat treatment of late aerial hyphae. The expression of the inducible hsp70 gene after heat shock also varied with the state of development and showed the highest inducibility in late aerial hyphae. Surface mycelium, from which aerial hyphae emerge, showed a similar increase in the amounts of both mRNA species. A developmental mutant (acon-2), which is defective in minor constriction budding of aerial hyphae, showed lower levels of con-2 mRNA as well as of 9-78 and hsp70 mRNA (after heat shock) in late aerial hyphae. The acon-2 mutant did not form conidia at this stage. It is concluded that the high constitutive and inducible expression of stress genes in late aerial hyphae is due to a developmental activation of their transcription or, alternatively, to a lower degradation rate of their mRNA during this stage.

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Purification and Partial Amino Acid Sequence of the Major 70,000-Dalton Heat Shock Protein in Neurospora crassa

Cited by 15 publications

References 0 publications

The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member

The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member

Temperature Adaptation of House Keeping and Heat Shock Gene Expression inNeurospora crassa

Differential expression of glucose-regulated (grp78) and heat-shock-inducible (hsp70) genes during asexual development of Neurospora crassa

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