As a consequence, hierarchical relationships, which require lunging behavior, were established in a smaller percentage of the fights (67% at the permissive vs. 10% at the restrictive temperature, Table 1).
This behavior is in contrast to the corresponding yeast TS-clathrin which failed to trimerize at the permissive and non-permissive temperature [36].
In this system, the Gal4-induced expression is suppressed by a ubiquitously expressed Gal80ts protein at the permissive temperature (18°C), but not at the restrictive temperature (30°C).
At the permissive temperature, the mutants are defective in 55Fe incorporation into nascent Pri2 polypeptide and exhibit reduced loading of multiple pol-prim subunits onto early replication origins.
This indicates that the TS-clathrin heavy chain is functional at the permissive but compromised at the non-permissive temperature.
In cells expressing the degron-myc tagged Ubc9, UV induced Rpb1 sumoylation occurred normally at the permissive temperature but was undetectable at the non-permissive temperature (Fig. 2B).
These cells have a temperature sensitive mutation in TAF1 which functions normally at the permissive temperature, 32°C, but is inactivated at the non-permissive temperature, 39°C [45].
The mutants were grown at the permissive temperature (25°C), shifted to 36°C for four hours before return to the permissive temperature.
These cultures were then transferred to plates with an aluminium replicator, then incubated at the permissive, semi-permissive and non-permissive (25°C, 30°C or 35°C) temperatures respectively on glucose containing media, then growth was scored.
To address this, wild-type and TS-mutant cells were incubated for a short period at the permissive or non-permissive temperature and subsequently stained for clathrin.
