Bac­te­ria Ac­ti­vate Fun­gal Gene Clus­ters

by S. Mar­vin Fried­man

Fungi are no­to­ri­ous for their abil­ity to pro­duce a wide va­ri­ety of sec­ondary metabo­lites, in­clud­ing an­tibi­otics, statins, im­muno­sup­pres­sants, my­co­tox­ins, and oth­ers. A ver­i­ta­ble phar­ma­copeia. In­ter­est­ingly, many of the gene clus­ters in­volved in the biosyn­the­sis of these com­pounds are silent un­der nor­mal lab­o­ra­tory con­di­tions. In some cases, the syn­the­sis of these com­pounds is known to de­pend on a sym­bi­otic re­la­tion­ship with bac­te­ria. Pre­vi­ous re­search has shown that in­ti­mate phys­i­cal con­tact be­tween the model fun­gus As­pergillus nidu­lans and the soil bac­terium Strep­to­myces ra­pamycini­cus ac­ti­vates the fun­gal ors gene clus­ter that en­codes one of the fun­gal sec­ondary metabo­lites, orsellinic acid and its de­riv­a­tives. The ef­fect is im­pres­sive: gene ex­pres­sion is 5 or­ders of mag­ni­tude greater when the fun­gus and bac­terium are co-cul­ti­vated. Orsellinic acid is a polyke­tide, a large group of or­ganic com­pounds that in­clude an­tibi­otics such as ery­thromycin, tetra­cy­clines, and am­pho­tericins. Orsellinic acid has phar­ma­co­log­i­cal ac­tiv­i­ties, in­clud­ing rad­i­cal scav­eng­ing.

Orsellinic acid. Source

In fil­a­men­tous fungi, the reg­u­la­tion of sec­ondary me­tab­o­lism in­volves the post-trans­la­tional mod­i­fi­ca­tion of hi­s­tones. Nutz­mann and cowork­ers have now an­a­lyzed the ef­fect of sev­eral such epi­ge­netic mod­i­fiers on the syn­the­sis of orsellinic acid by A. nidu­lans co-in­cu­bated with S. ra­pamycini­cus. Adding an in­hibitor of hi­s­tone acetyl­trans­ferase (HAT), lecanoic acid,  blocked tran­scrip­tion of the orsA gene. On the other hand, a hi­s­tone deacety­lase in­hibitor, suberoy­lanilide hy­drox­amic acid, ac­ti­vated the orsA gene with­out the need for co-in­cu­ba­tion with S. ra­pamycini­cus.

Model for hi­s­tone acety­la­tion-me­di­ated sec­ondary metabo­lite gene clus­ter ac­ti­va­tion in A. nidu­lans by S. ra­pamycini­cus. Sec­ondary metabo­lite genes (or­ange) un­der non­in­duc­ing con­di­tions are char­ac­ter­ized by deacety­lated hi­s­tone H3. The in­ti­mate con­tact be­tween A. nidu­lans and S. ra­pamycini­cus leads to an in­creased acety­la­tion of hi­s­tone H3 cat­alyzed by the Saga/Ada com­plex. The mod­i­fi­ca­tion of H3K9 is spe­cific for the sec­ondary metabo­lite gene clus­ters (red), whereas H3K14 acety­la­tion is not specif­i­cally tar­geted. Hence, Saga/Ada trig­gers the ex­pres­sion of the ors genes and the for­ma­tion of orsellinic acid (1), and other sec­ondary metabo­lites: lecanoric acid (2), F‑9775A (3), and F‑9775B (4). Source

By genome min­ing, the au­thors found 40 genes in A. nidu­lans tha­ten­code pu­ta­tive HATs. For 36 of them they could make dele­tion mu­tants (dele­tion of the re­main­ing four was ap­par­ently lethal). Of these 36, the one lack­ing the gcnE gene was dra­mat­i­cally im­paired in its abil­ity to in­duce the ors genes af­ter co-cul­ti­va­tion with the strep­to­mycete. This gene en­codes a HAT whose or­tholog in Sac­cha­romyces cere­visiae is part of sev­eral mul­ti­sub­unit ac­ti­va­tor com­plexes, one of which (the Saga/Ada com­plex) is in­volved in hi­s­tone acety­la­tion and chro­matin re­struc­tur­ing.

In A. nidu­lans, theSaga/Ada com­plex is in­volved in the reg­u­la­tion of biosyn­the­sis of sec­ondary metabo­lites such as sterig­ma­to­cystin, ter­re­quinone, and peni­cillin. This com­plex acety­lates pre­dom­i­nantly lysines K9 and K14 of hi­s­tone 3 (H3). The au­thors mea­sured the de­gree of hi­s­tone acety­la­tion of both the sterig­ma­to­cystin and ter­re­quinone gene clus­ters by chro­matin im­muno­pre­cip­i­ta­tion (ChlP)using an­ti­bod­ies against both of these lysines. The in­creased acety­la­tion af­ter 48 hour co-in­cu­ba­tion cor­re­sponds to the time of ex­pres­sion of these genes. Acety­la­tion lev­els of K9/K14 are sig­nif­i­cantly de­creased in the Saga/Adadeletion mu­tant. Fur­ther ChIP ex­per­i­ments showed that the pro­duc­tion of sec­ondary metabo­lites by A. nidu­lans is as­so­ci­ated witha global in­crease in the acety­la­tion of ly­sine K14 in H3. Speci­ficity for the ac­ti­va­tion of clus­ter genes, how­ever, is linked to H3K9 acety­la­tion.

To con­clu­sively prove that Saga/Ada di­rectly cat­alyzes the ors clus­ter-spe­cific acety­la­tion, the au­thors pre­pared a tagged ver­sion of GcnE, one of the es­sen­tial core com­po­nents of the com­plex. Us­ing ChlP, they showed that tagged GcnE is specif­i­cally re­cruited to the ors clus­ter dur­ing co-cul­ti­va­tion of the fun­gus and the bac­terium. It is no­table that al­though all four sec­ondary metabo­lite gene clus­ters ex­am­ined in this re­port de­pend on H3 acety­la­tion by the A. nidu­lans Saga/Ada com­po­nents GcnE/AdaB, only the ors clus­ter is ac­ti­vated by bac­te­r­ial-fun­gal in­ter­ac­tion. It ap­pears that some gene clus­ters, such as those in­volved in peni­cillin or sterig­ma­to­cystin biosyn­the­sis, re­quire ad­di­tional, more spe­cific sig­nals  for full ac­ti­va­tion, some of which have al­ready been iden­ti­fied. In the case of orsellinic acid pro­duc­tion, chro­matin re­struc­tur­ing in­duced by the bac­terium seems suf­fi­cient for full ac­ti­va­tion.

Phys­i­cal in­ter­ac­tion of A. nidu­lans with S. ra­pamycini­cus. Scan­ning elec­tron mi­cro­graph show­ing the close con­tact be­tween the fil­a­men­tous bac­te­ria and fun­gal mycelia. Bar = 1 μm. Source

What kind of sig­nal is passed from the bac­terium to the fun­gus? As re­ported in a pre­vi­ous pa­per by some of the same re­searchers, the su­per­nate of the bac­te­r­ial cul­ture did not ac­ti­vate the polyke­tides genes, nor did cul­tur­ing when the two part­ners were sep­a­rated by a dial­y­sis tube. In co-cul­ture, the two or­gan­isms ap­pear to be in di­rect phys­i­cal con­tact. All of this sug­gests that a dif­fusible com­pound is not in­volved, but the na­ture of the sig­nal be­tween the two part­ners re­mains un­known.

Sev­eral in­ter­ac­tions be­tween bac­te­ria and fungi have been ob­served in na­ture, such as the sym­bi­otic re­la­tion­ship in lichens and the sym­bio­sis of in­tra­cel­lu­lar bac­te­ria in zy­gomycetes. The cur­rent re­port is the first de­scrip­tion of a fun­gal hi­s­tone acety­la­tion sys­tem that is trig­gered by a bac­terium. A fun­da­men­tal ques­tion raised by this phe­nom­e­non is whether this in­duc­tion rep­re­sents a type of stress re­sponse or if it in­volves crosstalk be­tween mi­croor­gan­isms be­long­ing to dif­fer­ent do­mains. In­traspecies and in­ter­species cell-to-cell com­mu­ni­ca­tion in bac­te­ria, e.g. quo­rum sens­ing, has been well stud­ied in re­cent years, but re­ports such as this of a bac­terium talk­ing to a fun­gus are rare. If proven valid, I won­der what lan­guage is spo­ken.

Ref­er­ence

Nütz­mann HW, Reyes-Dominguez Y, Scher­lach K, Schroeckh V, Horn F, Gacek A, Schü­mann J, Her­tweck C, Strauss J, Brakhage AA. (2011). Bac­te­ria-in­duced nat­ural prod­uct for­ma­tion in the fun­gus As­pergillus nidu­lans re­quires Saga/Ada-me­di­ated hi­s­tone acety­la­tion. Proc Natl Acad Scie USA, 108 (34), 14282−14287. PMID 21825172

Mar­vin is Pro­fes­sor Emer­i­tus in the De­part­ment of Bi­o­log­i­cal Sci­ences at Hunter Col­lege of CUNY in New York City, and an As­so­ciate Blog­ger for Small Things Con­sid­ered.