The article discusses the role of Escherichia coli curli operons in directing amyloid fiber formation. Curli are extracellular fibers produced by E. coli and Salmonella that are involved in colonization of surfaces and biofilm formation. Biochemical, biophysical, and imaging analyses showed that curli fibers are amyloid-like. The CsgA curlin subunit, when purified in the absence of the CsgB nucleator, adopts a soluble, unstructured form that upon prolonged incubation assembles into fibers indistinguishable from curli. In vivo, curli biogenesis depends on the nucleation-precipitation machinery requiring the CsgE and CsgF chaperone-like and nucleator proteins, respectively. Unlike eukaryotic amyloid formation, curli biogenesis is a productive pathway requiring a specific assembly machinery. Bacteria express various cell-surface proteinacious filaments that promote colonization, entry into host cells, DNA exchange, and biofilm formation. Curli are highly aggregated, extracellular fibers involved in colonization of inert surfaces and biofilm formation. Polymerized curli appear as 4- to 7-nm-wide fibers by negative-stain electron microscopy. Under high-resolution EM, curli appear as a tangled and amorphous matrix surrounding the bacteria. Curli were purified and analyzed by SDS-PAGE, revealing that they are rich in β-sheet secondary structure. Curli induce a spectral change in Congo red solution and cause a red shift, indicating their amyloid-like properties. Amyloid formation in eukaryotic cells is thought to be the result of a misguided protein-folding pathway. In contrast, E. coli has a specific nucleation-precipitation machinery encoded by the csgAB and csgDEFG operons to assemble curli. CsgB is thought to nucleate CsgA fibers. The csgDEFG genes encode CsgD, a FixJ-like transcriptional regulator, and three putative curli assembly factors. The lipoprotein CsgG localizes to the inner leaflet of the outer membrane and may serve as a curli assembly platform. Deletion mutants of csgF and csgE resulted in aberrant CR binding properties and reduced fiber formation. However, these mutants could still guide CsgA polymerization. CsgA is secreted in a soluble, assembly-competent form by a csgB− mutant and can be assembled on the surface of csgA− mutant bacteria. CsgA produced by these cells was assembly competent, indicating a nucleation defect. CsgF may work independently or in concert with CsgB to guide in vivo extracellular nucleation of CsgA. The study demonstrates that E. coli can produce extracellular amyloid-like fibers, increasing the recognized functional repertoire of amyloid fibers and providing a useful model system to study their formation. Understanding the regulation of curli subunit polymerizationThe article discusses the role of Escherichia coli curli operons in directing amyloid fiber formation. Curli are extracellular fibers produced by E. coli and Salmonella that are involved in colonization of surfaces and biofilm formation. Biochemical, biophysical, and imaging analyses showed that curli fibers are amyloid-like. The CsgA curlin subunit, when purified in the absence of the CsgB nucleator, adopts a soluble, unstructured form that upon prolonged incubation assembles into fibers indistinguishable from curli. In vivo, curli biogenesis depends on the nucleation-precipitation machinery requiring the CsgE and CsgF chaperone-like and nucleator proteins, respectively. Unlike eukaryotic amyloid formation, curli biogenesis is a productive pathway requiring a specific assembly machinery. Bacteria express various cell-surface proteinacious filaments that promote colonization, entry into host cells, DNA exchange, and biofilm formation. Curli are highly aggregated, extracellular fibers involved in colonization of inert surfaces and biofilm formation. Polymerized curli appear as 4- to 7-nm-wide fibers by negative-stain electron microscopy. Under high-resolution EM, curli appear as a tangled and amorphous matrix surrounding the bacteria. Curli were purified and analyzed by SDS-PAGE, revealing that they are rich in β-sheet secondary structure. Curli induce a spectral change in Congo red solution and cause a red shift, indicating their amyloid-like properties. Amyloid formation in eukaryotic cells is thought to be the result of a misguided protein-folding pathway. In contrast, E. coli has a specific nucleation-precipitation machinery encoded by the csgAB and csgDEFG operons to assemble curli. CsgB is thought to nucleate CsgA fibers. The csgDEFG genes encode CsgD, a FixJ-like transcriptional regulator, and three putative curli assembly factors. The lipoprotein CsgG localizes to the inner leaflet of the outer membrane and may serve as a curli assembly platform. Deletion mutants of csgF and csgE resulted in aberrant CR binding properties and reduced fiber formation. However, these mutants could still guide CsgA polymerization. CsgA is secreted in a soluble, assembly-competent form by a csgB− mutant and can be assembled on the surface of csgA− mutant bacteria. CsgA produced by these cells was assembly competent, indicating a nucleation defect. CsgF may work independently or in concert with CsgB to guide in vivo extracellular nucleation of CsgA. The study demonstrates that E. coli can produce extracellular amyloid-like fibers, increasing the recognized functional repertoire of amyloid fibers and providing a useful model system to study their formation. Understanding the regulation of curli subunit polymerization